Current status of targeted therapy in non-small cell lung cancer.

Tumor tissue biomarkers are the basis for targeted therapy in non-small cell lung cancer (NSCLC). These biomarkers either reflect the target of the specific drug or some factor that might abrogate the effect of the drug. These targeted drugs are small-molecule inhibitors of a specific tyrosine kinase or a monoclonal antibody against a specific receptor. In this review, tissue biomarkers in NSCLC and companion diagnostics will be described, followed by an overview of targeted therapy to EGF/EGFR, HER2, VEGF/VEGFR, ALK, KRAS, BRAF, MEK and MET and some of the newer potential targets. Recent initiatives include the Lung Cancer Matrix Network, the BATTLE trials and the Lung Cancer Mutation Consortium. There are a number of key clinical trials in targeted therapy in NSCLC which will be reporting during the next year.

Effect of prior steroid treatment on temporal artery biopsy findings in giant cell arteritis.

AIM: To examine the effect of up to 6 weeks of corticosteroid treatment on the positive temporal artery biopsy rate in giant cell arteritis (GCA). METHODS: Prospective comparative clinical study of 11 patients meeting the American College of Rheumatology criteria for diagnosis of GCA. Patients underwent temporal artery biopsy within 1 week, at 2-3 weeks, or after 4 weeks of corticosteroid treatment. RESULTS: Overall, nine of 11 (82%) patients had positive temporal artery biopsies. Six of seven (86%) biopsies performed after 4 or more weeks of steroid treatment were positive. CONCLUSION: Temporal artery biopsy is useful several weeks after institution of steroids.

Donor-acquired small cell lung cancer following pulmonary transplantation.

We describe a case of donor-acquired small cell lung cancer after pulmonary transplantation for cystic fibrosis. The recipient was an ex-smoker with minimal smoking history and had been abstinent for 20 years. At the time of death, the donor chest radiographic finding was normal. The recipient had multiple posttransplant bronchoscopies and a normal CT scan result at 4 months after transplantation. The recipient presented 13 months after transplantation with metastatic disease. He did not respond to chemotherapy and died shortly thereafter. Molecular genetic techniques revealed that the primary tumor and metastases were different to recipient tissues, confirming the donor origin.

Hibernating myocardium: morphological correlates of inotropic stimulation and glucose uptake.

BACKGROUND: In patients with postischaemic left ventricular dysfunction, segments recovering function after revascularisation (hibernating myocardium) may not respond during dobutamine echocardiography, despite preserved [(18)F] 2-fluoro-2-deoxy-D-glucose (FDG) uptake at positron emission tomography. OBJECTIVE: To investigate whether this lack of response might reflect the degree of ultrastructural change in hibernating myocardium. METHODS: Transmural biopsies were obtained from 22 dysfunctional segments in 22 patients during coronary artery bypass grafting and examined by light and electron microscopy. Wall motion scores and coronary vasodilator reserve were assessed before and after coronary artery bypass grafting (CABG). RESULTS: Mean (SD) wall motion score improved in all segments following CABG (from 2.24 (0.4) to 1.55 (0.4); p < 0.0001), confirming hibernating myocardium. In these segments myocardial blood flow (positron emission tomography with H(2)(15)O) before CABG was similar to that in normal volunteers (1.02 (0.24) v 1.02 (0.23) ml/min/g), while the coronary vasodilator reserve was blunted (1.26 (0.7) v 3.2 (1.6); p < 0.0001). Myocardial blood flow was unchanged after CABG, whereas coronary vasodilator reserve increased to 2.10 (0.90) (p < 0.0007). In hibernating myocardium myofibrillar loss, interstitial fibrosis, and glycogen-rich myocytes were more marked than in control donor hearts. On the basis of the response to dobutamine before CABG, two functional groups were identified: group A, segments with inotropic reserve (n = 15); group B, segments without inotropic reserve (n = 7). FDG uptake was similar in group A and group B (0.40 (0.1) v 0.44 (0.1) micromol/min/g). In group B there was more myofibrillar loss (26 (8)% v 11 (5)%; p = 0.0009) and glycogen-rich myocytes (28 (11)% v 17 (10)%; p = 0.02), whereas interstitial fibrosis, myocardial blood flow, and coronary vasodilator reserve were similar in the two groups. Myofibrillar loss was the only independent predictor of inotropic reserve (p = 0.01). CONCLUSIONS: Hibernating myocardium is characterised by a reduced coronary vasodilator reserve which improves on revascularisation and shows a spectrum of ultrastructural changes that influence the response to dobutamine, while FDG uptake is invariably preserved.

Endoglin, an ancillary TGFbeta receptor, is required for extraembryonic angiogenesis and plays a key role in heart development.

Endoglin (CD105) is expressed on the surface of endothelial and haematopoietic cells in mammals and binds TGFbeta isoforms 1 and 3 in combination with the signaling complex of TGFbeta receptors types I and II. Endoglin expression increases during angiogenesis, wound healing, and inflammation, all of which are associated with TGFbeta signaling and alterations in vascular structure. The importance of endoglin for normal vascular architecture is further indicated by the association of mutations in the endoglin gene with the inherited disorder Hereditary Haemorrhagic Telangiectasia Type 1 (HHT1), a disease characterised by bleeding from vascular malformations. In order to study the role of endoglin in vivo in more detail and to work toward developing an animal model of HHT1, we have derived mice that carry a targeted nonsense mutation in the endoglin gene. Studies on these mice have revealed that endoglin is essential for early development. Embryos homozygous for the endoglin mutation fail to progress beyond 10.5 days postcoitum and fail to form mature blood vessels in the yolk sac. This phenotype is remarkably similar to that of the TGFbeta1 and the TGFbeta receptor II knockout mice, indicating that endoglin is needed in vivo for TGFbeta1 signaling during extraembryonic vascular development. In addition, we have observed cardiac defects in homozygous endoglin-deficient embryos, suggesting endoglin also plays a role in cardiogenesis. We anticipate that heterozygous mice will ultimately serve as a useful disease model for HHT1, as some individuals have dilated and fragile blood vessels similar to vascular malformations seen in HHT patients.

Tricuspid valve repair in a mountaineer.

Lord Hunt of Everest presented with aortic stenosis but predominant right ventricular failure. He was found to have signs of pulmonary hypertension with a dilated right ventricle, severe tricuspid regurgitation, and right atrial hypertrophy in the absence of elevated pulmonary artery pressures. He is a lifelong mountaineer and we attribute these findings to intermittent prolonged exposure to high altitude.

Unrestricted usage of immunoglobulin heavy chain genes in B cells infiltrating the wall of atherosclerotic abdominal aortic aneurysms.

The aim of this study was to provide evidence for the hypothesis that the B cell rich infiltrate concentrated in the adventitia of atherosclerotic abdominal aortic aneurysms is an autoimmune response to specific tissue antigens. Detailed histological examination of biopsies from 26 atherosclerotic abdominal aortic aneurysms showed in the adventitia, the presence of lymphoid follicles in 7/26 (27%) and of plasma cells in all cases. DNA prepared from the outer aneurysm wall (n = 25) was amplified using the polymerase chain reaction to investigate the repertoire of the immunoglobulin heavy chain (VH) genes used. Amplification of the VDJ region of VH, using both framework 2 and 3 primers, revealed unrestricted usage of the VH gene in 24/25 cases. The only case where restricted usage of the VH genes was observed, might have been attributable to severe virally-induced tissue inflammation. These results indicate that, in the vast majority of atherosclerotic abdominal aortic aneurysms, the B cell rich adventitial infiltrates are not an autoimmune response to a limited repertoire of tissue antigens.

Inducible nitric oxide synthase is present within human atherosclerotic lesions and promotes the formation and activity of peroxynitrite.

Inflammatory cytokines associated with atherosclerosis may be capable of stimulating the synthesis and activity of inducible nitric oxide synthase (iNOS), which could further influence the pathologic features associated with the disease. Although there is a certain amount of indirect evidence to support the presence of iNOS in atherosclerosis, there has been no definitive study to confirm this. This study has assessed the localization of iNOS within human normal and atherosclerotic vessels by immunocytochemistry, Western blotting, and in situ hybridization. Further, activity of NO synthase has been assessed by detection of nitrotyrosine, which is a marker indicative of the formation and activity of the nitric oxide-derived oxidant, peroxynitrite. In Western blots of crude homogenates of atherosclerotic aorta, the iNOS antiserum reacted with a band of approximately 130 kd (the known molecular weight for iNOS), but no such band was seen in normal aorta. Immunostaining and in situ hybridization confirmed the presence of iNOS in atherosclerotic vessels, in which it was specifically localized to (CD68-positive) macrophages, foam cells, and the vascular smooth muscle. The antiserum to nitrotyrosine reacted with a wide range of protein bands (approximately 180 to 30 kd) in Western blots of atherosclerotic aorta. The distribution of immunostaining for nitrotyrosine was virtually identical to that seen for iNOS and was present in macrophages, foam cells, and the vascular smooth muscle. In conclusion, these studies have demonstrated that stimulated expression of iNOS is associated with atherosclerosis and that the activity of this enzyme under such conditions preferentially promotes the formation and activity of peroxynitrite. This may be important in the pathology of atherosclerosis, which contributes to lipid peroxidation and to vascular damage.

Cytokine gene expression in aortic adventitial inflammation associated with advanced atherosclerosis (chronic periaortitis).

AIMS: To determine whether aortic adventitial chronic inflammation associated with advanced atherosclerosis ("chronic periaortitis") is associated with any detectable cytokine gene expression. METHODS: RNA was extracted from six fresh surgical specimens of atherosclertic aortic aneurysm wall showing a spectrum of chronic periaortitis. Controls included four normal aortas and an HUT 78 T cell line. Reverse transcriptase and the polymerase chain reaction (PCR) were used to amplify mRNA for interleukins-1 alpha (IL-1 alpha), -2 (IL-2), -4 (IL-4), IL-2 receptor-alpha (IL-2R-alpha), tumour necrosis factor alpha (TNF-alpha) and gamma interferon (IFN-gamma) with beta-actin as an internal control. RESULTS: No TNF-alpha mRNA was detected in any of the inflamed aortic tissue samples, in contrast to the aortic T lymphocytes propagated in culture in IL-2 conditioned medium (aortic cultured T cells) and peripheral blood mononuclear cells from these patients. In contrast, IFN-gamma, IL-1 alpha, IL-2, IL-2 receptor and IL-4 PCR products were detected for each inflamed aortic tissue RNA sample with IFN-gamma mRNA expression increasing with increasing degrees of adventitial inflammation. Only beta-actin mRNA was present in the normal aorta. CONCLUSIONS: These findings indicate the active nature of aortic adventitial chronic inflammation associated with human advanced atherosclerosis ("chronic periaortitis") and show its possible progressive potential to the clinically important diseases termed "idiopathic retroperitoneal fibrosis" and "inflammatory aneurysm".

The arteritides.

This article reviews the histopathological, clinical and immunological features of the arteritides. Based on these criteria, a classification scheme is proposed that includes infectious and non-infectious causes. Included in the non-infectious arteritides are: hypersensitivity vasculitis including serum sickness. Henoch-Schönlein purpura, mixed cryoglobulinaemia, hypocomplementaemia, drug and malignancy-associated vasculitis; arteritides of small and medium-sized arteries including polyarteritis nodosa, Kawasaki's disease, Wegener's granulomatosis, Churg-Strauss syndrome, necrotizing sarcoid granulomatosis, thromboangiitis obliterans (Buerger's disease) and localized forms of arteritis; arteritides involving large, medium and small-sized arteries which includes giant cell (temporal) arteritis, Takayasu's disease and arteritis of collagen-vascular disease (rheumatoid arthritis, rheumatic fever, Behçet's disease, Sjörgren's syndrome, systemic lupus erythematosis and systemic sclerosis.

The distribution of adhesion molecules in chronic periaortitis.

Chronic periaortitis is a local complication of human atherosclerosis. It is defined as the triad of advanced atherosclerosis, medical thinning and aortic adventitial chronic inflammation. It is present to a variable degree in association with atherosclerotic abdominal aortic aneurysms. These aortic adventitial infiltrates differ from those described solely within the atheroma itself, in that they consist predominantly of B lymphocytes. Many of the lymphocytes are activated and proliferating, and germinal centres are common. In this study, an immunohistochemical analysis was carried out on fresh surgical aortic aneurysm tissue in order to investigate the presence and distribution of activation-inducible adhesion molecules, and to correlate this with the degree of inflammation. A consistent finding was the presence of E-selectin on endothelial cells in up to 50% of the vessels throughout the aortic wall and at the base of the atheroma, independent of the severity of inflammation. ICAM-1 expression was abundant on many cell types and increased with the severity of chronic inflammation, being strongest in the germinal centres. VCAM-1 expression was predominant on follicular dendritic cells and also increased with severity of inflammation. VCAM-1 expression was also detected on vessels within lymphoid follicles. The pattern of expression of the adhesion molecules suggests a role in the initiation and progression of chronic inflammation associated with advanced atherosclerosis.

Quantification of specific mRNA by flatbed scintillation counting of dual-labeled dot blots.

A dual-labeling technique was developed for direct quantification of specific mRNA using a flatbed liquid scintillation counter. This method simultaneously measures cpm of 32P- and 35S-labeled probes bound to RNA dot blots and subtracts counts due to nonspecific background radioactivity bound to the filter. Probes for T-cell receptor and beta-actin (as the internal standard) were hybridized both separately and simultaneously to RNA isolated from five different sources. There was concordance between the radioactivity measured from single- and dual-hybridizations for each combination of 35S- and 32P-labeled probes. This methodology directly quantifies specific mRNA sequences bound to membranes and has potential for measuring gene dosage, without the need for re-probing or densitometric analysis.

The distribution of adhesion molecules in human atherosclerosis.

Chronic inflammatory cells are a recognized component of atherosclerotic plaques at all stages of development. As adhesion molecules play a fundamental role in inflammatory processes, we have carried out an immunohistochemical investigation of the distribution of endothelial leucocyte adhesion molecule-1 (ELAM-1)*, intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) in human atherosclerotic lesions. Autopsy specimens from abdominal aorta and coronary arteries were obtained from 21 cases within 24 h of death. ELAM-1 and ICAM-1 were consistently expressed by the entire intimal endothelium of normal coronary arteries and also by the intimal endothelium overlying aortic fatty streaks. Both coronary artery and aortic lesions showed strong staining for ICAM-1 on and around macrophages. VCAM-1 was not detected on intimal endothelial cells, but strong staining of adventitial lymphoid aggregates for this molecule was seen. This work suggests a role for ELAM-1 and ICAM-1 in mononuclear cell recruitment during atherogenesis.

Combined immunohistochemical and immunofluorescence method to determine the phenotype of proliferating cell populations.

AIMS: To determine the phenotype of proliferating cell populations. METHODS: The double immunostaining technique combines the autofluorescent properties of alkaline phosphatase substrate naphthol/Fast Red with immunofluorescence using fluorescein. Fresh human tonsil and fresh atherosclerotic aortic aneurysm wall tissue were studied using a panel of monoclonal antibodies including Ki-67, CD4, CD8, CD19, CD22, HLA-DR alpha, CD68 and CD31. RESULTS: This double immunostaining method permitted simultaneous colocalisation of different markers on the same cell and could be used to identify HLA-DR positive cells as well as proliferation associated Ki-67 positive cells in human tonsil tissue and in chronic periaortitis associated with advanced atherosclerosis. CONCLUSION: This technique is simple and the results may be viewed using a single fluorescence filter. The Fast Red reaction product is stable and does not fade under storage. The staining works particularly well with markers for nuclear antigens in combination with markers for cytoplasmic or surface antigens.

Fatal case of disseminated infection with the turtle bacillus Mycobacterium chelonae.

An apparently immunocompetent 78 year old woman presented with confusion, subcutaneous abscesses, and lesions of the nasopharynx. Gram positive, acid fast bacilli were isolated from her blood after 10 days' incubation. She was treated with trimethoprim-sulphamethoxazole for presumed disseminated nocardiasis but deteriorated and died. A post mortem examination showed skin and pulmonary lesions and endomyocardial fibrous plaques. Organisms isolated from the skin and lung were indistinguishable from those cultured from the blood. The organism was subsequently identified as Mycobacterium chelonae. Primary pulmonary infection and disseminated disease are rarely caused by this organism and bacteraemia is seldom documented. The clinical presentation and bacteriological and histological findings are difficult to differentiate from those of disseminated nocardiasis. Isolation of the organism may fail without prolonged incubation of initial cultures and there is a danger of its being dismissed as medically unimportant. Diagnosis is further hampered because large pulmonary foci may be poorly revealed by conventional radiological examination of the chest.

Storage of donor long saphenous vein.

Donor homograft vein is sometimes used in vascular surgery when autograft vein is not available. The optimum mode of storage remains controversial. An ideal solution would cause cellular disruption and thus decrease the immunogenicity of the donor vein, and allow a preserved collagen matrix and basement membrane, required to maintain the structure. Human donor vein was stored in normal saline at 4 degrees C, glycerol at 4 degrees C, liquid nitrogen and at -50 degrees C. The veins were examined at 1, 3, 7, 14, 21, 28, 35, 42, 49 and 56 days of storage using light microscopy, histochemistry, transmission and scanning electron microscopy. Vein stored at 4 degrees C in normal saline fulfilled the two requirements of preservation of the basement membrane and collagen matrix of the vein with loss of the cellular elements. We conclude, that in morphological terms this is the best mode of preservation. However further in vitro and in vivo studies are necessary.