Ewing in 1921 first described a distinct malignant tumour occurring in the soft tissues of the skeleton and called it “endothelial myeloma”. It is described in the literature as a relatively rare disease. Our records show a total of 27 cases of Ewing's tumour between the years 1941 to 1947. Of the total number of 9985 patients examined at the Tata Memorial Hospital between the period 1941 to April, 1945, 9640 were found to suffer from malignant disease of various tissues of the body. Sixty-five of these were primary malignant bone tumours: sixteen of these primary bone malignancies were Ewing's sarcoma. Thus of the total number of 27 Ewing's tumours seen to date, 16 were upto 1945 and 11 were seen between 1945 to the end of 1947. Tables The disease is more common in men than in women. Twenty of 27 patients were males and 7 females. Although no age seems to be exempt, the disease usually affects persons in the second and third decades of life. Nineteen of our patients were within this age period. The youngest was years of age, the oldest being 50. The duration of disease in all our cases, except two, was within 1 year. In one patient, the duration of symptoms was three and a half years.The bones of the pelvic girdle and the lower extremities are the most common sites of disease. It may, however, affect any part of the skeleton, including the vertebrae, ribs, skull and even the small bones of the hands and feet. In 19 of our cases the disease was located in the pelvic girdle and the lower extremities. None of our series had involvement of the vertebrae or the small bones of the extremities. When occurring in long bones, the disease was believed to affect only the shafts. In our material, the ends of long bones were often involved. Thus we had three cases with involvement of the ends of the femora, two in the upper end of the humerus, two in the upper end of the tibia and one in lower end of the fibula. In one patient, the lower end of the femur with the corresponding upper end of the tibia was involved . The disClinically, it usually presents with the well-known inflammatory complex of fever, pain, swelling and leucocytosis. A history of fever is not present in all. Pain is a constant feature. It varies from moderate to very severe, is often intermittent in character, and is more marked at night. Pain in orther parts of the skeleton is the earliest indication of metastasis. With this early metastasis the radiogram is usually negative. Six of our patients gave a definite history of trauma. The rest had either no trauma or no definite awareness of it. Trauma seems to have no direct causal relation to bone tumours, for were it so the incidence of bone tumours should be much higher than it actually is, and yet it is difficult to escape the impression that trauma in some cases is an exciting cause. Enlargement of the nearest draining lymph nodes is often present. Five of our patients had enlarged nodes, three in the groin, one in the neck and one in the popliteal fossa. Two of our patients (Nos. 1230 & 6545) had rounded masses in the upper abdomen. In one of these, the bone lesion was in the upper tibia, and the other in the mid-femoral region. The abdominal masses were noticed two and five months respectively after the bone lesion was seen. In both these cases, the possibility of a primary  neuroblastoma cannot be excluded. The disease progresses fairly rapidly metastasizing rapidly to the nearest lymph nodes, the lungs and the distant bones and leading to a fatal termination with-in a few months to a couple of years.All of our patients, except one, showed leucocytosis, the white blood cell (WBC) count varying between 8 to 20 thousands. One patient had a WBC count of 4000. Seven patients had moderate to severe anaemia, the red blood cell (RBC) count being below 4 millions on admission. In the rest, the count was about 4.5 millions. The serum calcium varied round 10 mg%; the phosphorus values were within normal limits. Alkaline phosphatase varied between 1.5 to 5.5 B Units. Acid phosphatase values were normal. Kahn's test was negative in 18 patients; in the rest, the test was not done.In osteogenic sarcoma the values for serum alkaline phosphatase were between 10 to 16 B Units. The higher values of alkaline phosphatase in osteogenic sarcoma could be utilised as an additional diagnostic help for differentiation from Ewing's sarcoma.Roentgen examination of bone tumours is now established as an indispensable diagnostic procedure. For some time, fixed roentgen criteria were ascribed to different varieties of tumours. For instance the “sun-ray” appearance was considered characteristics of osteogenic sarcoma, the “onion peel” appearance for Ewing's tumour , “soap bOsteolytic destruction of the involved bone is the most frequent radiological finding . Twenty-Condensation of bone, when present may occur intracortically or subperiosteally . IntracoA variety of patterns is presented by sub-periosteal new bone formation. The well known “onion peel” appearance is only occasionally seen in Ewing's tumour. On the other hand, it is sometimes seen in chronic pyogenic or syphilitic osteomyelitis. Its presence therefore does not confirm nor its absence exclude the diagnosis of Ewing's tumour. Sometimes the subperiosteal new bone is laid down in thin irregular striations placed perpendicular to the shaft, resembling the “sun-ray” appearance , thoughtFrom the radiogram alone, a differentiation of Ewing's tumour has to be made from a variety of bone lesions. These include osteomyelitis , osteolyIt is now widely accepted that a positive diagnosis of a malignant bone lesion requires the combined efforts of a surgeon, the radiologist and the pathologist. The pathologist is obviously the final arbiter. And yet often the pathologist does not arrive at a positive diagnosis from his microscopic picture alone without reference to the skiagram and the clinical picture. Histologic diagnosis is not written on the slide. The microscopic appearance has to be interpreted. Leaving aside the personal factor, then the microscopic appearance, particularly with Ewing's tumour admits of different interpretation. A fair amount of polemical literature has collected about the nature of this bone lesion. Although most experienced oncologists following Ewing, accept this lesion as a distinct disease process, an emphatic opposition to such a view is voiced by some pathologist of reputed e.g, Willis, Colville, and Steruburg. They regard all cases of Ewing as “precocious” metastatic bone lesions from an occult neuroblastoma either in the adrenal or the sympathetic ganglia. However Willis in his latest book “Pathology of Tumours” published in 1948, says “some cases at least of Ewing's syndrome are due to clinically obstructive metastasis of neuroblastoma in long bones”(p. 864). One could ill afford to hold a dogmatic opinion on this count. The only course open is that of taking the “majority” view as a working hypothesis, always keeping in mind to look for data which go to disprove such hypothesis.The cytological appearance of Ewing's tumour is fairly constant. It shows “sheets” or groups of cells, with varying amount of fibrous strands in between. The cells are usually round with some variation in size. Cell walls are indistinct with scanty cytoplasm and large hyperchromatic nuclei. Nuclei have well-defined margins with scattered chromatic material. Sometimes the cells are polygonal when closely packed. Cells may be arranged along blood vessels and the RBCs may be separated from the tumour cells only by endothelium and delicate strands of reticulum, the so-called perithelial arrangement.Occasionally the cells are arranged in a “pseudo-rosette” formation. These are structures with a circular arrangement of cells, the nuclei of the cells being situated at their outer poles. Sometimes argyrophil reticulum fibres are seen scattered between groups of cells.These microscopic appearances form the basis of a diagnosis of “Ewing's Tumour”. This picture differs from the original description of the cell type by Ewing. He described it as “a small polyhedral cell with pale cytoplasm, a small hyperchromatic nucleus, and a well-defined cell wall.” More recent reports (Stout and Jaffe) however described cells to have a large nucleus, scanty cytoplasm and indistinct cell walls.Jaffe and Leichtenstein do not ascribe any diagnostic significance to the “perithelial” arrangement of cells. They state that haemorrhage into the neoplastic tissue is accompanied by ingrowth of blood vessels. Many of these vessels becomes surrounded by tumour cells. According to them, the “vessel spaces are not lined by tumour cells.” Besides, they state that a similar perivascular arrangement of tumour cells is seen in other sarcomas when focal areas have undergone extensive haemorrhage.The “pseudo-rosette” appearance in Ewing's tumour is cause of much controversy. Similar structures are seen during the development of the sympathetic nervous system and frequently in neuroblastomas arising from the adrenals and sympathetic system. Proper staining of these however will show neurites protruding from the central ends of the cells. The central cores will show fibrillar or granular appearances. Jaffe and Leichtenstein state that the cores of the so-called rosette arrangement in Ewing's tumour do not show a fibrillar or granular appearance, but are made up of degenerated nuclei and necrosis, the radiating arrangement of cells being explained on a purely physical basis. Stout, himself is not satisfied with this explanation. He says ‘orientation of the nuclei towards the peripheral pole has a teleological aspect which suggests an activity other than pure degeneration. Consequently, their presence in the Ewing's tumour remains for me unexplained and in need of further elucidation”Colville and Willis (1933) and Willis (1940) because of close resemblance of the cytologic picture of Ewing's tumour with metastasis from sympathetic neuroblastoma, warned against making a diagnosis of Ewing's tumour from clinical and biopsy data. Only carefully executed autopsies can exclude neuroblastoma. Without this a diagnosis of Ewing's sarcoma is not justified.A perusal of the available literature on the subject shows that most experienced oncologists  while accepting the weighty objection of Willis et al are inclined to regard Ewing's sarcoma as a disease entirely separate from metastatic bone lesions from sympathetic neuroblastoma, despite absence of autopsy examination in most of their cases and on the basis of the biopsy microscopic picture alone. Other lesions which often require to be differentiated from Ewing's tumour are reticulum cell sarcoma, lymphocytic lymphoma, and Hodgkin's disease. Sometimes bone metastasis from a silent highly malignant carcinoma may be the source of error, when miscroscopically it presents a uniform pattern of round cells.Ewing's tumour, like reticulum cell and lipogenic sarcoma, is highly radio-sensitive. High radio-sensitivity in Ewing's tumour means a rapid initial regression of the tumour mass and not a complete control of the malignant process nor even in most cases an extended survival. With this high sensitivity of the tumour, theoretically it should be possible to obtain an extended survival if not complete cure by early adequate irradiation. Yet few “cures” are reported. The tumour is highly malignant and rapidly spreads to distant parts. The disease generally extends more along the medulla than is shown by the radiogram, so that treatment of the tumour area as shown by the radiogram would be inadequate. A large portion of the adjacent normal bone should be included in the field.In most cases this would require inclusion in the field of the whole of the bone. A tumour doze of 4500r at least is necessary. Woodard and Coley have recently reported that a tissue dose of 3000 to 5000r could control the tumour for varying period; after this dosage a large number of viable tumour cells were still present. They report complete destruction of all tumour cells with tumour dose approaching 6000 to 7000r. Two of their patients so treated are living after eleven years. This shows that despite its apparent sensitivity, for complete control the tumour requires doses as high as does a squamous cell tumour. This would necessarily entitle much damage to the normal bone. It is not generally realised that adult normal bone is more vulnerable to irradiation than surface epithelium. Besides it is now definitely established that absorption of penetrating radiations by bone is higher than in soft tissues. Very high tumour doses necessary to control Ewing's tumour will permanently damage the adjacent normal bone which has to be included in the treatment field.th to the 6th week dense new bone becomes definitely established. The irregular cortical margins are smoothened and layers of dense bone are deposited along the shaft which may show hardly any evidence of the malignant process.Under irradiation the initial regression of Ewing's tumour is often amazingly rapid, with control of pain and marked reduction in soft tissue swelling . There iAll case of Ewing's tumour do not show this high sensitivity. Some are definitely resistant. Two of our patients gave very poor response to irradiation. One of these, an eleven year old girl with involvement of the upper tibial metaphysis had no reduction in pain or swelling. The patient died within two months with lung metastases. An opportunity was afforded to us for noticing this radioresistance while treating two almost identical cases referred to us from the British Military Hospital in Bombay. The right ilium was the seat of disease in both, with nearly similar roentgen appearance. Both were treated during the same period with identical deep therapy technique. One of these showed excellent response. Bone regeneration was seen within four weeks. The second patient had no reduction in pain. Soft tissue swelling was slightly reduced and the patient gradually went down during treatment.Our results of radiation treatment so far as extended survival is concerned are extremely disappointment. Twenty-two patients came up for X-Ray treatment. All of these except one, were treated with 200 kV. 0.5 mm Cu plus 1 mm. AL filter, 50 cm F.S.D; and one was treated with 400 kV. 4 mm Cu plus 1 mm. AL filter 50 cms. distance. The doses ranged from 2400r (air) on each of the ports in cases of long bones, to 3000 and 3500r on each of two ports in flat bones. Three of the treated cases were seen alive without metastases for 1 year; after that there was no “follow up”. Three were seen up to 6 months without disease; then no follow-up. Six patients never came for follow-up after completion of treatment. One lived for two years but with metastases in various parts of the body. We chased the disease from place to place with X-ray treatment, each time with excellent symptomatic improvement, till finally, the patient succumbed to extensive metastases in the spine. Three patients, treated within the last four mouth of 1947 are still living - two without metastases and one with metastases.


Tumour blood flow was characterised in a 'tissue-isolated' rat tumour model, in which the vascular supply is derived from a single artery and vein. Tumours were perfused in situ and blood flow was calculated from simultaneous measurement of (1) venous outflow from the tumour and (2) uptake into the tumour of radiolabelled iodo-antipyrine (IAP). Comparison of results from the two measurements enabled assessment of the amount of blood 'shunted' through the tumours with minimal exchange between blood and tissue. Kinetics of IAP uptake were also used to determine the apparent volume of distribution (VDapp) for the tracer and the equilibrium tissue-blood partition coefficient (lambda). lambda was also measured by in vitro techniques and checks were made for binding and metabolism of IAP using high-pressure liquid chromatography. VDapp and lambda were used to calculate the perfused fraction  of the tumours. Tumour blood flow, as measured by IAP (TBFIAP), was 94.8 +/- 4.4% of the blood flow as measured by venous outflow, indicating only a small amount of non-exchanging flow. This level of shunting is lower than some previous estimates in which the percentage tumour entrapment of microspheres was used. The unperfused fraction ranged from 0 to 20% of the tumour volume in the majority of tumours. This could be due to tumour necrosis and/or acutely ischaemic tumour regions. For practical purposes, measurement of the total venous outflow of tumours is a reasonable measure of exchangeable tumour blood flow in this system and allows for on-line measurements. Tracer methods can be used to obtain additional information on the distribution of blood flow within tumours.


P<0.0001). LYVE-1/PCAB detected LVI in 25/67 cases (37.3%) and their presence was significantly associated with both lymph node metastasis  and unfavourable overall survival (OS) (P=0.0453), while not relapse- free survival (RFS) (P=0.2948). LMVD had no influence for RFS and OS . Our study demonstrates that immunohistochemistry with LYVE-1/PCAB is a highly sensitive method for detecting tumour LV/LVI in breast cancer and LVI is a useful prognostic indicator for lymphatic tumour dissemination.This study was undertaken to determine the highly sensitive method for detecting tumour lymphatic vessels in all the fields of each slide (LV), lymphatic microvessel density (LMVD) and lymphatic vessel invasion (LVI) and to compare them with other prognostic parameters using immunohistochemical staining with polyclonal (PCAB) and monoclonal antibodies (MCAB) to the lymphatic vessel endothelial hyaluronan receptor-1 (LYVE-1), and the pan-endothelial marker factorVIII in a series of 67 human breast cancers. In all LYVE-1-stained sections, LV (some of which contained red blood cells) were frequently found localised in extralobular stroma, dermis, connective tissue stroma and adjacent to artery and vein, but were rare within the intralobular stroma or the tumour body (3/67 cases) or areas of widespread invasion. In contrast small blood vessels were observed in intra- and extralobular stroma in the factor VIII-stained sections. Quantitation of vessel numbers revealed that LYVE-1/PCAB detected a significantly larger number of LV than either H&E or LYVE-1/MCAB ( It has been well established that angiogenesis is necessary for tumour growth and haematogenous metastasis  had to have primary, unilateral, breast cancer and no other malignancy. Clinical and pathological data are listed in μm thick sections stained with haematoxylin and eosin (H&E) were assessed histopathologically and were used to select the maximal area of all the cut surfaces of the tumour that included the invasive components. Immunostains for FVIII-related antigen were performed on paraffin sections using the streptavidin-biotin-immunoperoxidase method as previously described  and overall survival (OS) by fitting Kaplan–Meier survival curves . For comparison of number of lymphatic vessel assessed by the three different staining methods, for association of LMVD and clinical or pathologic parameters and for the association of LVI and lymph-node status, Kruskal–Wallis test, Mann–Whitney l curves  to varioBoth the polyclonal and monoclonal anti LYVE-1 antibodies yielded specific and consistent staining of endothelial cells in the lymphatic vessels . Many lyP<0.0001). Strong significant correlation was between the LYVE-1/PCAB-immunostained lymphatic vessels and LYVE-1/MCAB-immunostained lymphatic vessels . Median LMVD was 6.1 microvessels mm−2 (range 0–17.9 vessels). A strong significant correlation was found between LMVD and LYVE-1/PCAB-immunostained lymphatic vessels . There was no significant correlation between the LMVD and BMVD . An inverse correlation was seen between histological grading and LMVD (P=0.0434), while histological grading or menopausal status trended with the number of lymphatic vessels (P=0.0712) or LMVD (P=0.0944). There was no significant correlation between clinical tumour size, lymph-node status, LVI, or estrogen receptor and LMVD or the mean number of lymphatic vessels , but LVI detected by LYVE-1/PCAB was significantly associated with lymph-node status , while no significant difference in RFS (P=0.2948). However, LMVD had no influence for OS and RFS .The mean and median (range) number of all lymphatic vessels is shown in  vessels . LVI wasThis manuscript has focused on the utility of different LYVE-1 antibodies as routine markers for detecting and quantitating lymphatic vessels in breast cancer. Our results confirm that both LYVE-1 polyclonal and monoclonal antibodies distinguish efficiently between lymphatic and blood vessels in pathological specimens. However, due to the greater dependence of LYVE-1 monoclonal antibodies on tissue fixation and antigen retrieval methods that result in partial antigen destruction, we have found that LYVE-1 polyclonal antibodies are suited to routine immunohistochemical staining applications. Furthermore, using immunohistochemical staining with LYVE-1 polyclonal antibodies we have shown that lymphatic invasion is positively associated with lymph node involvement and unfavorable OS.Quantitation of tumour lymphatic vessels for the purpose of tumour staging has for decades been problematic. Although morphology can sometimes distinguish lymphatic vessels from blood vessels by the frequent absence of a basement membrane and lack of erythrocytes in the latter, neither is a reliable method for routine use. These considerations have hampered the reliable identification of tumour lymphatic vessels in routine histopathology. More recently, however, the development of specific markers such as the lymphatic hyaluronan receptor LYVE-1, the subject of this manuscript, has allowed many new experimental studies of tumour lymphatics to be initiated. To date, the majority of these studies have employed LYVE-1 polyclonal antibodies, requiring microwave treatment or pressure cooking for antigen retrieval . HoweverPrevious studies have reported that all blood vessels that contain erythrocytes are negative for LYVE-1 supporting its specificity for lymphatics . A promiSeveral recent studies in animal models have reported that lymphatic vessels are frequently observed in the peripheral rim of the tumour, but not in the body of the tumour itself . Most stRecent studies have demonstrated that VEGFR-3 or D2-40 immunostained microvessels was associated with either node metastases (The current study has used LYVE-1 staining to increase the accuracy and rate of detection of LVI since using this method makes it easy to distinguish from BVI . In rece


The rat Walker 256 tumour was grown in mice that had previously been thymectomized and treated with anti-lymphocyte serum. These rat tumour-bearing mice were used to determine the therapeutic indices of 4 anti-tumour drugs.The agent with the highest index of the four examined was 5-aziridino 2,4-dinitrobenzamide (CB1954), followed by melphalan, aniline mustard and methotrexate, in that order. This rank order is the same as that found when therapeutic indices are determined on the Walker tumour growing in the rat. In this system, therefore, drugs have been ranked correctly in effectiveness against a rat tumour by measuring their effects on the tumour when growing in an immunosuppressed xenogeneic species. The implications for testing the drug sensitivity of individual human tumours before treating the patient are discussed.


We examined hepatic artery ligation combined with an angiogenesis inhibitor, TNP-470, in the treatment of VX2 tumour inoculated into the liver of rabbits. Effects on tumour growth were correlated with arterial collateral development in this system. Three treatment methods were compared: (1) the left hepatic artery was ligated at the liver hilum (ligation group); (2) TNP-470 (40 mg per body) was infused continuously for 7 days via the common hepatic artery (TNP group); (3) the left hepatic artery was ligated and TNP-470 was infused continuously for 7 days via the common hepatic artery (ligation + TNP group). These treatments were started 12-14 days after tumour inoculation. The day of initiating treatment was defined as day 0. Although there were no significant differences in tumour volume among the three treated groups on day 7 after treatment, tumour volumes in the ligation + TNP group were significantly smaller than in the ligation group and the TNP group on day 14 after treatment. The vasculature and arterial collaterals around the tumour were demonstrated by the perfusion of a silicon rubber solution, Microfil. In the ligation + TNP group, the new microvasculature around the tumour decreased compared with the ligation group. The TNP-470 inhibition of microvascular proliferation may limit the development of collaterals that communicate with new feeding arteries. These results suggest that transarterial embolization combined with TNP-470 may enhance the anti-tumour effect of transarterial embolization alone in the treatment of liver tumours.


In this study, we have undertaken a detailed analysis of expression of the TIMP family in normal human brain and malignant gliomas at both the mRNA and protein level. Reverse transcription-PCR (RT-PCR) analyses of total RNA from surgical tumour specimens revealed unique expression patterns for the 4 members of the TIMP family, with TIMP-1 and -4 showing positive and negative correlations, respectively, with glioma malignancy. By RT-PCR, TIMP-2 and TIMP-3 expression did not change with tumour grade. In situ hybridization localized TIMP-1 to glial tumour cells and also to the surrounding tumour vasculature. TIMP-4 transcripts were predominantly localized to tumour cells, though minor expression was found in vessels. Recombinant TIMP-4 reduced invasion of U251 glioma cells through Matrigel, and U87 clones overexpressing TIMP-4 showed reduced invasive capacity in vitro. TIMP-4, but not TIMP-1, blocked Membrane Type-1-MMP-mediated progelatinase-A (MMP-2) activation in human umbilical vein endothelial cells. The differential expression and localization of individual TIMPs may contribute to the pathophysiology of human malignant gliomas, particularly with regard to tumour vascularization. © 2001 Cancer Research Campaign


Growth of isogeneic transplants of a spontaneous murine adenocarcinoma, which is virtually devoid of tumour-specific transplantation antigens, is inhibited by i.v. injection of C. parvum 3 days after tumour inoculation, or by mixing a small dose of C. parvum with the tumour inoculum. Moreover, the therapeutic effect of cyclophosphamide, followed by i.v. or i.p. injection of C. parvum 5 days later, on established transplants of the same tumour is greater than that of cyclophosphamide alone. These findings are consistent with the hypothesis that in both situations  the effect of C. parvum is largely due to activation of macrophages or macrophage precursors. They have the important practical implication that adjuvant therapy with C. parvum may be of value, even with tumours which are devoid of TSTA.


The introduction of the “maximum tolerated dose” in usual treatment protocols (and its concomitant overt toxicity) made necessary the imposition of rest periods between cycles of therapy—a practice that not only involves re-growth of tumour cells, but also growth of selected clones resistant to the therapy. To avoid the problems caused by traditional chemotherapeutic regimens, a new modality of drug administration called “metronomic chemotherapy” has been proposed. This name makes reference to the chronic, equally spaced administration of  low doses of various chemotherapeutic drugs without extended rest periods. The novelty of this treatment modality lies not only in its antitumour efficacy with very low toxicity, but also in a cell target switch, now aiming at tumour endothelial cells. The knowledge acquired in the experimental field of metronomic chemotherapy, plus the increasing experience that is being obtained in the clinical setting, will help to lead a change in the design of therapeutic protocols against cancer. The introduction of the maximum tolerated dose (mtd) in usual treatment protocols made necessary the imposition of rest periods between cycles of therapy—a practice that not only involves re-growth of tumour cells, but also growth of selected clones resistant to the therapy. Hence, the therapeutic success obtained during the first cycles of treatment reverts in the direction of growth of more malignant metastatic tumours with no therapeutic response.Chemotherapy regimens reflect a controversy that is by now historical: between efficacy in tumour killing and lack of toxicity, which way should the scale be tipped? On one side is the ability of chemotherapeutic drugs to disrupt the A turning point in cancer chemotherapy can be placed in the year 2000, when Fidler and Ellis said, “Cancer is a chronic disease and should be treated like other chronic diseases” mct) built its foundations with plenty of experimental work, beginning with the pioneering work in the Folkman and Kerbel laboratories. Browder and colleagues demonstrated that standard chemotherapeutic drugs such as cyclophosphamide can also be used as anti-angiogenic agents. The administration of cyclophosphamide in doses lower than the mtd, at shorter intervals and without extended rest periods, showed results better than those obtained with the mtd schedule in the treatment of two cyclophosphamide- resistant tumours, Lewis lung carcinoma and the murine mammary carcinoma cell line EMT-6 mtd, the dose of cyclophosphamide was still high, and the experimental animals received palliative care to ameliorate gastrointestinal dysfunction and weight loss.As with any other experimental therapy, metronomic chemotherapy . In the in vivo model, animals bearing xenografts of both neuroblastoma cell lines were treated with low doses of vinblastine; or a monoclonal neutralizing antibody (DC101) that blocks the function of vascular endothelial growth factor receptor 2 (vegfr2) and, hence, vascular endothelial growth factor (vegf) itself; or both agents together. The animals treated with either drug alone showed inhibition of tumour growth, an effect that did not last. On the other hand, the combination treatment induced an initial response similar to that in the other treatment groups, and that response was followed by long-term tumour regression. The usual signs of drug toxicity in mice  were not seen at the doses used in the experiment in vitro testing of the effect of non-cytotoxic doses on endothelial cell functions involved in angiogenesis and then, in vivo, with the chick embryo chorioallantoic membrane model. They showed dose-dependent anti-angiogenic activity At almost the same time, Klement These seminal experiments suggested the possibility that various kinds of tumours may be amenable to chronic treatment, prompting our group, and others, to study the efficacy of metronomic chemotherapy in other tumour models. We focused our interest both on the antitumour and anti-metastatic effect achieved, and on the toxicity of the treatment. We were able to demonstrate that metronomic administration of cyclophosphamide at low doses on a 3-times-weekly schedule eradicated established rat lymphomas and sarcomas (10 mg/kg and 5 mg/kg body weight respectively). Neither metastatic growth nor recurrence at primary sites occurred for 100% of the lymphomas and 83% of the sarcomas, and no weight loss was observed. At the same time, the regimen was devoid of hematologic, cardiac, hepatic, and renal toxicity mct in various in vivo tumour models in which animals were treated metronomically with diverse agents. Established orthotopic multidrug-resistant human breast cancer xenografts in scid (severe combined immunodeficiency) mice were treated with paclitaxel, vinblastine, cisplatin, or doxorubicin alone or in combination with DC101 scid mice vegfr2 inhibitor (SU5416) or two different inhibitors of matrix metalloproteinases uft (a 5-fluorouracil pro-drug) showed therapeutic efficacy in a model of advanced metastatic breast cancer mtd regimens mtd in a so-called chemo-switch regimen, suggesting a potential clinical strategy in which standard mtd is followed by a novel maintenance regimen The experimental work developed in several laboratories showed antitumour efficacy and a lack of the manifest toxicity of tor SU541 or two dMore recently, a new therapeutic approach was proposed, now targeting the multicellular biologic entity of the tumour microenvironment. Blansfield and colleagues vegf and basic fibroblast growth factor (bfgf) exceed those of angiogenesis inhibitors such as thrombospondin-1 (tsp-1) Angiogenesis is a normal process that has essential roles in development, reproduction, and tissue repair. On the other hand, pathologic angiogenesis is closely involved in diabetic retinopathy, chronic inflammation, and tumour formation. Angiogenesis plays a critical role in the growth and metastatic spread of tumours In 1971, Judah Folkman first articulated the concept of what he called “anti-angiogenic” drugs. Based on observations that expansion of a tumour mass was limited in the absence of angiogenesis, he proposed that treatment with drugs that prevent the formation of tumour blood vessels might be able to constrain cancer for prolonged periods vegf is detected in many malignant tumours in vitro and in vivo studies demonstrated that vegf can be considered a marker of the angiogenesis process vegf was observed in patients with advanced breast cancer treated with metronomic low-dose cyclophosphamide vegf levels during treatment The angiogenic polypeptide mct is the fact that the most effective dose and schedule have yet to be defined. Therefore, with the aim of filling that need, the roles of various surrogate markers of angiogenesis and vasculogenesis were studied to understand whether they can be used to define the anti-vascular activity of certain drugs or drug combinations, and to predict response or survival in cancer patients receiving anti-angiogenic treatment vegf and tsp-1 might act as surrogate markers for monitoring treatment activity ii study of mct with etoposide and cyclophosphamide in combination with daily thalidomide and celecoxib for recurrent malignant gliomas in adults, the serum and urine levels of vegf, bfgf, endostatin, and tsp-1 were evaluated. Nonsignificant differences between responders and non-responders were found et al., who studied vegf serum concentrations in patients with advanced breast cancer receiving metronomic low-dose oral cyclophosphamide and methotrexate plus or minus thalidomide, obtained opposing results. Compared with baseline levels, mean levels of vegf 2 months after treatment were significantly lower vegf was studied as a prognostic factor in patients with recurrent or metastatic squamous cell carcinoma of the head and neck treated with metronomic paclitaxel. Preliminary results suggested an association between vegf and both response and disease stabilization One of the drawbacks for the extended clinical use of cecs) are seldom found in the blood of healthy individuals . However, cecs are detected in patients with neoplastic, inflammatory, and vascular conditions ceps) that are considered to be an alternative source for some of the endothelial cells of newly formed blood vessels represent a subset of immature vegfr2- positive cecs also found in peripheral blood A number of recent experimental observations suggest that the growth of some types of cancer may depend on vasculogenesis  and not just angiogenesis . Circulating endothelial cells  used alone or in combination with an anti-angiogenic drug Several ceps was reported to be usable as a surrogate marker for angiogenesis and, thus, as a biomarker for monitoring targeted anti-angiogenic drug activity ceps can be considered to be the optimal biologic dose cecs and their precursors in patients with relapsed or refractory aggressive non-Hodgkin lymphoma, suggesting that this combination may be working by inhibiting angiogenesis, a result that should be validated in a larger patient sample cecs and clinical outcome in patients with advanced breast cancer receiving mct. They found that cecs were a particularly good predictor of disease-free and overall survival after a prolonged follow-up of more than 2 years cecs and ceps. In accord with that result, no significant clinical activity was demonstrated. Therefore, it is possible that changes in circulating cecs and ceps could be observed in the setting of more effective anti-angiogenic therapies The level of et al. demonstrated that low-dose cyclophosphamide inhibits tumour growth and induces selective apoptosis of endothelial cells within the tumour vascular bed by up-regulating the endogenous angiogenesis inhibitor tsp-1. As compared with wild-type tumours, tumours treated with low-dose cyclophosphamide showed similar expression of both matrix metalloproteinases and basement membrane– derived angiogenesis inhibitors; tsp-1 expression, on the other hand, was significantly increased et al. reported that protracted in vitro exposure of endothelial cells to low concentrations of several anticancer agents, including microtubule inhibitors and an alkylating agent, caused marked induction of tsp-1 gene and protein expression. Conversely, an increase was detected in circulating tsp-1 in plasma of scid mice bearing PC-3 human prostate cancer treated with metronomic low-dose chemotherapy tsp-1–null mice tsp-1 can induce apoptosis of microvascular endothelial cells expressing CD36 In vitro and in vivo experiments showed that mct induces the expression of tsp-1 and that this up-regulation has a negative effect on endothelial cell survival tsp-1 as a secondary mediator of the anti-angiogenic effect of at least some mct regimens.Hamano mct with certain chemotherapeutic drugs could be the stimulation of the immune response, because metronomic administration of oral cyclophosphamide in advanced cancer patients induces a profound and selective reduction in circulating regulatory T cells (Tregs). This effect is associated with suppression of Treg inhibitory functions on conventional T and natural killer cells, leading to restoration of peripheral T-cell proliferation and innate killing activities mct with cyclophosphamide. Euthymic rats and nude mice bearing a rat B-cell lymphoma were treated metronomically with cyclophosphamide. After a short period of tumour growth, all treated rats showed sustained tumour regression, an effect that none of the non-treated control rats showed. Interestingly, neither treated nor control nude mice showed tumour regression, a result that could be ascribed to the absence of the adaptive immune response Another mechanism responsible for the antitumour effect of ceps) and escape from the immune response. The antitumour and anti-metastatic effects of metronomic chemotherapy would be achieved through several mechanisms, including inhibition of ceps, anti-angiogenic activity, and depending on the tumour and the drug or drugs administered, direct cytotoxic action on tumour cells or stimulation of the immune response, or both.mct is emerging in the era of targeted cancer treatment. To date, several articles have been published reporting the use of mct in phase i or ii trials with drugs such as cyclophosphamide, doxorubicin, capecitabine, thalidomide, 5′-deoxy-5-fluorouridine, etoposide, paclitaxel, methotrexate, and prednisone, among others, with encouraging results.The novel therapeutic approach of mct in a clinical setting was published in 2002. Colleoni and colleagues evaluated the clinical efficacy of low-dose methotrexate and cyclophosphamide in heavily pretreated breast cancer patients, obtaining significant efficacy with minimal toxicity mct is worthy of further clinical evaluation.The first work using et al. performed a pharmacoeconomic evaluation of metronomic cyclophosphamide –methotrexate and a number of other novel phase ii regimens for the palliative treatment of metastatic breast cancer and concluded that the mct scheme could reduce health care costs, especially those associated with the combined use of the new highly expensive molecularly targeted therapies In an interesting study, Bocci mct regimens are administered, are supporting mct implementation in the clinical setting with a predicted result of increased survival and life quality.The preliminary evidence of disease stabilization obtained in patients with varying and progressing tumours, and the low toxicity profile registered when mtd, which induces important remissions, is generally followed by recurrences with the development of tumours even more malignant.The data so far obtained induced us and other authors to begin a changing of our way of thinking about cancer treatment. To date, the aim of chemotherapy has been to achieve complete tumour suppression, a goal reached only exceptionally. Tumour elimination has been elusive, and metastasis accounts for an important part of this failure. We now know that all tumour cells cannot be consistently eliminated by high dosing schemes. The repeated administration of We can now focus on cancer therapy from a different angle. With low-dose chemotherapy, it may be possible to obtain a therapeutic effect in the clinical setting similar to that obtained experimentally. Should this hypothesis prove true, great progress will have made toward the solution of the cancer problem. On the other hand, present knowledge of tumour biology creates the recognition that tumour presence is not incompatible with patient survival. Therefore, it might not be necessary to aim for complete tumour eradication. A change from the apparently remote therapeutic objective of killing all tumour cells to the more pragmatic objective of diminishing tumour burden as much as possible and maintaining that diminishment over time can now be considered. This goal might be achieved by administering drugs in a low-dose metronomic schedule over time.mct, plus the increasing experience that is being obtained in the clinical setting, will spark a change in the way in which basic and clinical researchers design their therapeutic protocols against cancer.It is foreseeable that the knowledge acquired in the experimental field of


However, attempted sensitization and subsequent challenge with DNCB demonstrated that the afferent limb of the responses was impaired in some patients. This did not appear to be related to the morphology of the tumour or to prognosis.Defects in cellular immunocompetence have been sought in 25 patients with Kaposi's sarcoma. Skin tests with recall antigens, and PHA lymphocyte stimulation in vivo and in vitro and these correlated significantly with the morphology and histology. The interpretation of the results for an individual is confounded by the multiplicity of factors influencing the outcome in a particular patient.Tumour specific reactions were demonstrated both


At a cutoff value of 4 U ml−1, the sensitivity was 73%. TNM and Dukes' classification of the tumours revealed a strong correlation between faecal Tumour M2-PK levels and staging. The determination of Tumour M2-PK in faeces provides a new promising screening tool for colorectal tumours.Proliferating cells, especially tumour cells, express a special isoenzyme of pyruvate kinase, termed M2-PK, which can occur in a tetrameric form with a high affinity to its substrate, phosphoenolpyruvate (PEP), and in a dimeric form with a low PEP affinity. In tumour cells, the dimeric form is usually predominant and is therefore termed Tumour M2-PK. The levels of Tumour M2-PK within tumours and in EDTA-plasma correlate with staging and the ability of the tumour cells to metastasise. Since most colorectal tumours grow intraluminally, it appeared interesting to determine whether Tumour M2-PK is detectable in the faeces of tumour patients. Stool samples were tested by ELISA from controls without colorectal cancer and colorectal cancer patients. Whereas Tumour M2-PK levels were low in the control group (mean value±s.e.m.: 3.3±0.4,  Tumour M2-PK. The tetramer : dimer ratio of M2-PK can be quantified by gel permeation, isoelectric focusing or by ELISA  . Whereas M2-PK.) . In gastby ELISA . The uprby ELISA . The tetnd A-Raf  . For sample preparation, 100 mg of the single, random stool specimen was extracted in 10 ml extraction buffer and diluted 1 : 10 in the sample washing buffer provided. The ELISA plate is coated with a monoclonal antibody against Tumour M2-PK. Tumour M2-PK from stool samples or standards binds to the antibody. A second monoclonal antibody, which is biotinylated, binds to Tumour M2-PK during the next incubation. Both monoclonal antibodies against Tumour M2-PK specifically react with Tumour M2-PK (dimeric form of M2-PK) and do not crossreact with the other isoforms of pyruvate kinase . In the next incubation step, a conjugate of peroxidase (POD) and streptavidin binds to the biotin moiety. POD oxidises the substrate 3,3′,5,5′-tetra-methyl benzidine (TMB) and the oxidised form of TMB is determined photometrically at 450 nm.For the statistical comparison of the nonparametrically distributed data, the Kruskall–Wallis ANOVA test was used .Sensitivity was calculated as the number of patients with colorectal cancer who tested positive for Tumour M2-PK (true positives) divided by the total number of histologically confirmed tumour patients , expressed as a percentage (i.e. sensitivity=(number of true positives/number of tumour patients) × 100).Specificity was calculated as the number of controls who tested negative for Tumour M2-PK (true negatives) divided by the total number of control subjects , expressed as a percentage  × 100).−1, the sensitivity was calculated to be 73% and the specificity as 78%. TNM classification of the colorectal tumours according to the criteria of the American Joint Committee on Cancer , giving an average coefficient of variance (CV) of 7.9% (3.5–13.6%). The interassay variance was calculated with five samples between 4 and 73 U ml−1, tested on 10 different days. The average CV was 7.3% (3.8–12.6%).The intra-assay variance was evaluated by 18-fold determination of five samples  revealed that increased Tumour M2-PK in tumour cells is a general metabolic alteration during tumorigenesis and correlated with malignancies of the tumours  gene were not detected in any stool samples of healthy controls . Furthermore, the overexpression of a mutated M2-PK protein suppresses A-Raf transformation in NIH 3T3 cells, which underlines the central role of M2-PK within the tumour metabolome (glutaminolysis (http://www.metabolic-database.com).In contrast to many other genes, no mutations have yet been found in the M2-PK gene (tabolome . The tetinolysis  (http://The oscillation between the highly active tetrameric form and the nearly inactive dimeric form allows optimal adaptation of the tumour metabolome to the varying nutrient and oxygen supply constantly found in solid tumours (Tumour M2-PK is detected with an ELISA that can be easily performed in every routine laboratory. Samples are stable and the sensitivity of the Tumour M2-PK test is higher than the detection of mutations in p53, ras or APC (


Acceleration of secondary tumour growth and metastases following excision of a primary tumour has been attributed to the consequent removal of primary tumour-generated inhibitory factors. However, our studies have shown that surgical wounding of normal tissues significantly stimulated the growth of malignant tissues without the concomitant presence or excision of a tumour mass. A humoral stimulating component was indicated by the proliferative response of tumours and metastases distant from the surgical wound. All 16 human and murine tumours, of nine different histologies, showed a measurable acceleration of growth when implanted in surgically treated animals, suggesting that the ability of malignant tissue to respond to surgical wounding of normal tissue was not histologically or species specific. The proliferative surge of malignant tissues was detectable soon after wounding and had a duration of 2-3 days. The surgical wound as the source of the tumour-stimulating factor(s) was affirmed by the significant inhibition of tumour proliferative responses when a somatostatin analogue was applied topically to the surgical wound within 1 h of wounding, and/or during the critical tumour-stimulatory period of 1-2 days after wounding. A potential therapeutic window for reducing a risk factor that may be inadvertently imposed upon every surgical/oncology patient is indicated.


The anti-tumour activity of CPT-11, a topoisomerase I inhibitor, was evaluated in four human neural-crest-derived paediatric tumour xenografts; one peripheral primitive neuroectodermal tumour (pPNET) (SK-N-MC) and three neuroblastomas. Two models, SK-N-MC and IGR-N835, were established in athymic mice from a previously established in vitro cell line. Two new neuroblastoma xenograft models, IGR-NB3 and IGR-NB8, were derived from previously untreated non-metastatic neuroblastomas. They exhibited the classic histological features of immature neuroblastoma along with N-myc amplification, paradiploidy, chromosome 1p deletions and overexpression of the human mdr 1 gene. These tumour markers have been shown to be poor prognostic factors in children treated for neuroblastoma. CPT-11 was tested against advanced stage subcutaneous tumours. CPT-11 was administered i.v. using an intermittent (q4d x 3) and a daily x 5 schedule. The optimal dosage and schedule was 40 mg kg-1 daily for 5 days. At this highest non-toxic dose, CPT-11 induced 100% tumour-free survivors on day 121 in mice bearing the pPNET SK-N-MC xenograft. For the three neuroblastoma xenografts, 38-100% complete tumour regressions were observed with a tumour growth delay from 38 to 42 days, and anti-tumour activity was clearly sustained at a lower dosage (27 mg kg-1 day-1). The efficacy of five anti-cancer drugs commonly used in paediatric oncology or in clinical development was evaluated against SK-N-MC and IGR-N835. The sensitivity of these two xenografts to cyclophosphamide, thiotepa and cisplatin was of the same order of magnitude as that of CPT-11, but they were refractory to etoposide and taxol. In conclusion, CPT-11 demonstrated significant activity against pPNET and neuroblastoma xenografts. Further clinical development of CPT-11 in paediatric oncology is warranted.


Jun-Yang Liou, is a co-corresponding author of this paper with Kenneth K. Wu. His e-mail address is


The levels of deoxythymidine kinase in tumour cells (C-TK) and in serum (S-TK) were investigated and the tumour volume calculated in 89 patients with non-Hodgkin lymphoma (NHL), in order to ascertain the importance of C-TK and tumour burden as regards the S-TK levels. Among all patients, a correlation was seen between S-TK and tumour volume but not between S-TK and C-TK. However, within different tumour volume categories , there was a correlation between S-TK and C-TK. Multiple regression analysis supported this notion. C-TK correlated with the proliferation-associated parameters, S-phase fraction and mitotic index. As already known, S-TK was found to have a strong prognostic value. C-TK and tumour burden were also of prognostic value. In multivariate analyses, C-TK and tumour volume did not provide prognostic information in addition to S-TK, whereas, in the absence of S-TK, C-TK and tumour volume did provide additional information. It is concluded that the serum level of TK depends on both the tumour burden and the tumour cell proliferation rate. Based upon estimations of S-TK in patients assessed shortly after chemotherapy, we also suggest that S-TK reflects the number of proliferating cells that have died during the period immediately before sampling.


Experiments were conducted to determine the dose response of rat bladder urothelium to a range of different single and fractionated intravesicular doses of the carcinogen, N-methyl-N-nitrosourea (MNU). A dose-related response of bladder-tumour incidence to single graded doses of MNU was found, and a threshold does suitable for use of multistage carcinogenesis experiments was derived from these data. For any given total dose of MNU, the tumour incidence was greater if the MNU had been administered in several small fractions than if it had been administered in fewer larger ones. Extending the interval between doses did not reduce the tumour incidence. It is argued that these results support the multistage theory of carcinogenesis. The histopathology and cell-surface alterations which characterize the development of MNU-induced bladder cancer are described and the contribution of hyperplasia and calculi are discussed.


A 4-day tumour sensitivity assay of potential use in predicting tumour response to cytotoxic drugs has been investigated in patients with chronic lymphocytic leukaemia. The method comprised isolation of white cells from peripheral blood, drug exposure and incubation for 4 days. Drug-induced tumour cell kill was assessed by differential staining of dead and live cells such that the latter could be morphologically identified, with subsequent calculation of tumour cell viability. Concentrations of drug for use in the assay were chosen for chlorambucil (2 micrograms ml-1), 4-hydroperoxy-cyclophosphamide (2 micrograms ml-1)--which was used in vitro in place of cyclophosphamide--prednisolone (0.5 microgram ml-1) and vincristine (0.1 microgram ml-1), to give a scatter of values which was in good agreement with clinical expectations. In 21 cases where the in vitro result could be compared with the in vivo response, there were 4 true positive comparisons , 15 true negative comparisons (resistant both in vitro and in vivo) and 2 false positive comparisons . A result was obtained in 86% (65/76) of samples received. The assay appears to show considerable promise as a tumour chemosensitivity test and warrants wider investigation, including prospective in vivo/in vitro correlations that could be based on the results presented here.


A micro-cytotoxicity assay was adapted for the detection of cell-dependent antibodies (CDA). Using normal rat spleens as the source of effector cells such CDA activity was readily demonstrable in allo-immune sera tested on cultured sarcoma cells. The same technique was then used to examine for tumour specific antibodies in the sera of Hooded rats bearing a “non-immunogenic” syngeneic metastasizing sarcoma. During the early stages of tumour growth, at Days 7 and 14, tumour specific CDA cytotoxicity was detectable at high titres. By Day 21, however, this activity had completely disappeared from the serum. This cell-dependent cytotoxicity was tumour specific in that it did not kill cells from an unrelated syngeneic sarcoma, and the activity was probably confined to immunoglobulin G as detected by molecular weight separation techniques. Following tumour amputation at Day 21, this type of specific antibody activity rapidly re-appeared in the serum. The presence of tumour specific CDA showed an inverse correlation with the presence of specific inhibitors of cell-mediated immunity in the same sera. At no stage in tumour growth could complement-dependent cytotoxicity be detected in tumour bearing rat sera.It is concluded that cell-dependent cytotoxic activity is not associated with conventional complement dependence, that this CDA type of assay is exquisitely sensitive and is suitable for the detection of anti-TSTA antibodies in tumour bearing rats.The possible significance of CDA activity in syngeneic tumour immunity is discussed briefly. The results suggest that the role of humoral immune mechanisms in host resistance to tumour growth needs re-appraisal.


The effects of intravenous diethylamine/nitric oxide (DEA/NO), a short-acting nitric oxide (NO) donor, on systemic haemodynamics, muscle and tumour blood flow (MBF and TBF) and tumour oxygenation were examined in rats bearing subcutaneous R3230Ac carcinoma in the leg. The effects of DEA/NO on the diameters of tumour-feeding and normal arterioles were evaluated in window chambers with and without implanted tumours. DEA/NO reduced mean arterial pressure (MAP) when given at doses > or = 100 nmol kg(-1), with maximal suppression at 0.5-1 min followed by return to baseline within 20 min. DEA/NO did not affect MBF except at the highest doses (500 and 1000 nmol kg(-1)). In contrast, DEA/NO reduced TBF and constricted tumour arterioles at doses > or = 100 nmol kg(-1). Tumour arteriolar vasomotion occurred in more than half the animals during hypotension and with a significantly higher frequency than in normal granulating tissue at a dose of 500 nmol kg(-1). Normal arterioles rapidly and significantly vasodilated for about 3 min and then returned to baseline. The reductions in TBF and MAP were accompanied by synchronous reduction in tumour pO2. Our findings suggest that DEA/NO decreases TBF in two ways. In the window chamber model, vascular steal occurs as normal arterioles adjacent to tumour dilate more than tumour arterioles during the initial period of hypotension. In leg tumours, the predominant mechanism is attributable to reduced perfusion pressure induced by lowered MAP, which decreases flow to the tumour, probably because of relatively higher flow resistance. The vasoconstriction and vasomotion in tumour arterioles during DEA/NO-induced hypotension may reflect differences in regulatory metabolism of NO between neoplastic and normal arterioles. Thus, intravenous injection of a short-acting NO donor, DEA/NO, decreases MAP and heart rate, leading to subsequent decreases in tumour blood flow and oxygenation.


Fluid overload, a common complication following cardiac surgery in infants is often difficult to manage. Dialysis can be used in severe cases, but may not be well tolerated. In such patients, peritoneal drainage could be an alternative option for fluid removal. We report the case of a newborn with a truncus arteriosus who developed postoperatively a complicated clinical course with right ventricular dysfunction, prerenal condition as well as fluid overload despite diuretic therapy. Dialysis was indicated for fluid removal. Peritoneal dialysis was started using a surgically placed Tenckhoff catheter and stopped due to inefficacy and leaks and no other modalities of dialysis were used. However, the catheter was left in place over a period of two months for fluid drainage and removed because of unexplained fever. In order to determine the effect of peritoneal drainage, we selected a period of one week before and one week after the removal of the drain to compare daily clinical data, urine electrolytes and renal function and found a positive effect on fluid balance control. We conclude that the fluid removal by continuous peritoneal drainage is a simple and safe alternative that can be used to control fluid balance in infants after cardiac surgery. Fluid overload is a common complication following cardiac surgery in infants. The cause is thought to be multifactorial and includes pre-existing cardiac dysfunction, exogenous fluid during cardiopulmonary bypass, postsurgery acute kidney injury and the strain on the heart caused by the surgical procedure –3. ConseThe patient described here was born to a mother known with insulin dependant type 2 diabetes. The pregnancy was overall uncomplicated, but antenatal ultrasounds revealed the presence of a truncus arteriosus. Delivery was induced at 40 weeks gestational age and was complicated by shoulder dystocia as well as thick meconium fluid, but no invasive resuscitation was required. Birth weight was 4.0 kg and Apgars were 7, 9, 9 at 1, 5 and 10 minutes, respectively.μmol/L, blood urea nitrogen of 2.2 mmol/L; urine analysis was normal as well as renal Doppler ultrasound. Preoperative weight was 3.9 kg. The corrective cardiac surgery was performed at 8 days of life. During extracorporeal circulation that lasted 198 minutes, a modified ultrafiltration used during 36 minutes drained 275 milliliters of fluid. The aortic cross was clamped during 60 minutes. At the end of the surgery, fluid balance was positive of 550 cc. During the follow-up, the patient had multiples episodes of worsening renal function and fluid retention despite diuretic therapy. It was a prerenal dysfunction with a fractional excretion of sodium of less than 1% consistent with a hemodynamic cause. The first episode of renal dysfunction began immediately after cardiac surgery. Serum creatinine concentration increased from a preoperative value of 30 μmol/L to 107 μmol/L on day 1 with a maximum of 120 μmol/L on day 4 and returned to normal with value of 30 μmol/L on day 11 after surgery. In parallel, the cardiac function remained difficult to stabilize after the surgery with a decrease in the contractility of the right ventricle noted on day 9 postsurgery. Three weeks postsurgery, the patient had remained on ventilatory support because of persistent cardiac failure mainly due to right ventricular dysfunction. On post-operative day 23, the serum creatinine concentration reached a peak of 105 μmol/L, and the patient's weight was 60% over his birth weight. Low volume peritoneal dialysis through a surgically placed cuffed Tenckhoff catheter was initiated with 10 mL/kg exchange volume and the dwell time was one hour. Glucose-based peritoneal dialysis solutions were used with concentration of glucose of 1.5% and 2.5%. After repetitive episodes of blood pressure drop following the drainage with incidental hypoxemia and difficulties with dialysate leaks, peritoneal dialysis had to be discontinued 3 days later. The peritoneal drain was kept in place and used for continuous drainage over a period of 2 months from day 23 to day 85 in an attempt to prevent or treat significant fluid accumulation and was removed because of an unexplained fever despite multiples investigations. Over this period, there were no signs of infection at entry site or peritonitis and peritoneal fluid cultures remained negative.Cardiac ultrasound confirmed the presence of only congenital abnormalities with a truncus arteriosus type I associated with atrial and ventricular septal defect. Preoperative renal function evaluation was normal with serum creatinine of 30 μmol/L). Although peritoneal drainage slightly improved the fluid balance, the clinical course of the patient worsened; he remained ventilated because of both cardiac and thoracic oedema and died on day 118 after surgery from decompensated cardiac failure, sepsis, pulmonary infection, and worsening renal function.In order to determine the effect of peritoneal drainage on fluid balance and diuresis, we analyzed the data from the week prior and following the discontinuation of the peritoneal drainage on day 85 . During We present the case of an infant in whom a continuous peritoneal drain was used successfully to assist the management of fluid overload after cardiac surgery without decreasing urine output nor impairing renal function. Fluid overload is a common complication following cardiac surgery in infants. Several treatments have been proposed to treat fluid overload in the context of cardiac failure and cardiac surgery including diuretics and peritoneal dialysis –5. Most Cardiac surgery in the newborn is often complicated by fluid overload. While peritoneal dialysis is needed to help regulate fluid balance when acute kidney injury is present, the continuous peritoneal drainage is a simple alternative technique to remove fluid in these patients when dialysis is not indicated. Further data collection from other similar patients is however needed to confirm this hypothesis.


Subcellular fractions from an aminoazo dye induced rat hepatoma (D23) were examined for their ability to evoke rejection responses in syngeneic hosts to transplanted tumour cells and to induce the production of humoral antibody. Membrane fractions isolated by zonal centrifugation and displaying an increased activity of tumour specific antigen , as well as crude membrane fractions and purified tumour cell ghosts, all elicited tumour specific antibody demonstrable by membrane immunofluorescence staining of viable hepatoma D23 cells. Tumour cell nuclei or soluble cytoplasmic protein were, however, lacking in this capacity. Resistance to tumour cell challenge was not observed in rats treated with any of the hepatoma D23 subcellular fractions administered by various routes either alone or in admixture with bacterial adjuvants. These findings are relevant to current views that tumour immunity may be more optimally achieved by inoculation of intact (viable or attenuated) tumour cells.


Serum from rats bearing progressively growing aminoazo dye-induced rat hepatomata has been fractionated by Sephadex G150 gel filtration chromatography and isolated fractions have been examined by indirect membrane immunofluorescence techniques to detect tumour specific antigen and antibody. Hepatoma D23-specific antigenic activity was associated with material  isolated in the included volume of the gel at pH 7·3. The fraction excluded from the gel  was adjusted to pH 3·0 and further separated by Sephadex G150 gel filtration chromatography at pH 3·0 into gel included and excluded fractions. Hepatoma D23 specific antibody, demonstrable by membrane immunofluorescence staining of hepatoma D23 cells, was found to be eluted in the excluded volume and specific antigenic activity was retarded into the included volume of the gel. These results indicate that hepatoma D23 bearer serum contains free circulating tumour specific antigen in excess, together with specific immune complexes. The presence of these factors in tumour bearer serum is discussed in terms of “blocking” phenomena whereby serum factors may protect tumour cells from sensitized lymphocyte cytotoxic attack.


Many factors have been individually related to outcome in populations of non-small-cell lung cancer (NSCLC) patients. Factors responsible for the outcome of an individual after surgical resection are poorly understood. We have examined the importance of 'tumour volume' in determining prognosis of patients following resection of NSCLC in a multivariate model. Cox's proportional hazard analysis was used to determine the relative prognostic significance of stage, patient age, gender, tumour cell-type, nodal score and estimated 'tumour volume' in 669 cases with NSCLC treated with surgical resection, of which 280 had died. All factors  were individually related to survival (P < 0.05). When examined together, survival time was significantly and independently related to 'tumour volume' and stage (P < 0.001), and other factors ceased to be significant. In cases with stage I or II tumours, risk of death was found to increase significantly with increasing estimated 'tumour volume' . In cases with stage IIIa tumours this factor alone was the significant prognostic variable. In conclusion, an estimate of 'tumour volume' significantly improves prediction of prognosis for individual NSCLC patients with UICC stage I or II tumours.


We have previously documented the development and subsequent disappearance of cytolytic activity mediated by lymphocytes from lymph nodes draining Moloney sarcomas destined either to regress or grow progressively. We now report that these tumour-draining lymphnode cells (LNC) that were no longer cytotoxic, spontaneously regenerated peak levels of killing after culture in vitro for 4 days in the absence of exogenous tumour antigen. Cytolytic activity, which was antigenically specific, was mediated by T lymphocytes. Resurgence of cytolytic activity in vitro was accompanied by proliferative changes  which peaked on the 3rd day of culture. Although normal, nonimmune LNC underwent quantitatively similar proliferative changes in culture, the killing that developed was weak and antigenically nonspecific. Transfer of cultured, tumour-draining LNC to immunologically compromised, syngeneic mice conferred complete protection from Moloney sarcoma progression. Adoptive transfer could be delayed for 6 days after tumour induction without loss of protection. These results suggest that there exists in Moloney sarcoma-bearing mice a mechanism that limits the differentiation of pre-killer cells into cytolytically active T lymphocytes, and that such inhibition is eliminated when LNC are explanted into culture.


Serum activity of alpha-N-acetylgalactosaminidase , the extracellular matrix-degrading enzyme that appears to be produced exclusively by cancer cells, was measured in mice bearing SCCVII tumours (squamous cell carcinoma). The NaGalase levels in these mice increased with time of tumour growth and were directly proportional to tumour burden. After exposure of SCCVII tumours to a single X-ray dose of 20 Gy, the serum NaGalase levels gradually decreased during the first 10 days after treatment  and then began to increase. The decrease in serum NaGalase activity was more rapid after the treatment of SCCVII and EMT6 tumours by photodynamic therapy (PDT) and was dependent on the PDT dose. The treatments (based on photosensitizers Photofrin or mTHPC) that were fully curative resulted in the reduction of NaGalase activity to background levels within 2 or 3 days after PDT. A slower decrease in NaGalase activity was found after PDT treatments that attain an initial tumour ablation but are not fully curative. The regrowth of PDT-treated SCCVII tumours was preceded by an increase in serum NaGalase levels, which was detected as early as 8 days before the visible tumour reappearance. These findings ascertain the validity of serum NaGalase measurement for the assessment of tumour response to different treatments and support the concept that the NaGalase measurement could serve as a diagnostic and prognostic index that might allow oncologists to design the dosage or nature of treatment.


D-myo-inositol-1,2,6-triphosphate  is a polyanionic molecule capable of chelating divalent metal ions with anti-tumour and anti-cachectic activity in a murine model.4.To investigate the role of zinc in this process, mice bearing cachexia-inducing MAC16 tumour were treated with AT, with or without concomitant administration of ZnSO−1, AT effectively attenuated both weight loss and growth of the MAC16 tumour, and both effects were attenuated by co-administration of Zn2+. The concentration of zinc in gastrocnemius muscle increased with increasing weight loss, whereas administration of AT decreased the levels of zinc in plasma, skeletal muscle and tumour, which were restored back to control values after administration of ZnSO4.At a dose of 40 mg kgThese results suggest that zinc is important in both tumour growth and cachexia in this animal model. Tumour fragments were obtained from donor animals selected as those having the maximum weight loss, and were implanted s.c. into the flank by means of a trochar, as previously described (−1) or AT (40 mg kg−1) plus 8.05 μg ZnSO4 in 25 μl PBS administered i.v. 1 h after AT. A fourth group received ZnSO4 alone. Both PBS and AT were administered s.c. three times a day. Tumour volume, body weight and food and water intake were monitored daily. Animals were killed by cardiac puncture under terminal anaesthesia when the body weight loss was at, or before, 20%, as approved by the British Home Office. The ethical guidelines that were followed met the standards required by the UKCCR guidelines , which were obtained from our own inbred colony, and were fed a rat and mouse breeding diet  and water M, slit 0.7 mm, using air/acetylene gas and a lamp current of 10 mA. Standards of known concentrations were used to construct calibration curves from which the concentration of zinc in the sample could be determined.Tissues were weighed in crucibles, which were previously washed in 5% Lipsol, and rinsed with deionised bio-filtered water, followed by trace analysis using nitric acid. The crucibles were placed in a furnace at room temperature, and the temperature was increased to 150°C for 1 h, then 600°C for 2 h, followed by 800°C overnight. A fine white ash, free from organic material, was obtained, and the crucibles were cooled and stored in desiccators. The crucibles were subsequently re-weighed to obtain the mass of ash, which was dissolved in 10 ml of trace element analysis concentrated nitric acid and sonicated for 30 min to ensure stabilisation. The dissolved ash was then transferred to a 100 ml volumetric flask, which was made up to volume with deionised bio-filtered water. The sample was then analysed for zinc using a PerkinElmer Analyst 100 atomic absorption spectrophotometer , at a wavelength of 213.9 nP-values <0.05 were considered significant.Results are presented as mean±s.e.m. Differences in means between groups were determined by one-way analysis of variance (ANOVA), followed by the Tukey–Kramer multiple comparison test. −1 of AT was chosen, as this has been shown to produce optimal inhibition of both weight loss and tumour volume  through phosphorylation of p65/Rel A at multiple serine residues (κB in the muscle of mice has been shown to lead to atrophy through increased expression of proteasome subunits and the E3 ligase, MURF1 (α-subunit, and increased protein degradation through the NF-κB-mediated induction of the ubiquitin–proteasome pathway (α and the activity and expression of the ubiquitin–proteasome pathway have been shown to be attenuated by AT in mice bearing the MAC16 tumour (The results of this study show the importance of Zn pathway . The act2+. Studies in vitro also support a separate role of AT in attenuating muscle atrophy. Thus, using murine myotubes as a model of skeletal muscle, AT has been shown to attenuate both the induction of protein degradation and depression of protein synthesis induced by proteolysis-inducing factor, angiotensin II, lipopolysaccharide and TNF-α (2+. These studies also suggest that Zn2+ is required for the caspase-3/-8-mediated activation of PKR. As AT is highly negatively charged, it is unlikely that it would readily penetrate into cells; hence, it is likely that the effects are mediated through the chelation of extracellular Zn2+.These results provide some evidence that the attenuation of loss of skeletal muscle by AT is separate from its anti-tumour activity, although both effects seem to be due to its ability to chelate Zn2+, and the role of zinc in the proliferation of pancreatic cancer (2+ alone attenuated tumour growth rate to the same extent as AT. However, it is known that although tumours need Zn2+ to grow and survive, excess Zn may induce apoptosis (2+ were shown to stimulate cell proliferation through activation of the mitogen-activated protein kinase and phosphatidylinositol-3-kinase pathways mediated by a Gq-coupled receptor. (2+ led to desensitisation of the zinc receptor possibly through constitutive desensitisation, as for other G-protein-coupled receptors, and through inhibition of tumour growth. These results could explain the apparent anomaly that both chelators of Zn2+ and Zn2+ itself inhibit tumour growth.In view of the potential mechanism by which AT attenuates tumour growth rate by chelation of Znpoptosis . Extrace2+. Further studies are required on the role of Zn2+ in the signalling cascade leading to muscle atrophy, in order to elucidate the mode of action of this important metal in relation to cachexia.These results suggest that AT is a novel agent with anti-cachectic and anti-tumour activity due to its ability to chelate Zn


Selective apoptosis of endothelial cells was detected in vivo in sections of Colon 38 tumours in mice within 30 min of administration of 5,6-Dimethylxanthenone-4-acetic acid (25 mg kg−1). TUNEL staining intensified with time and after 3 h, necrosis of adjacent tumour tissue was observed. Apoptosis of central vessels in splenic white pulp was also detected in tumour-bearing mice but not in mice without tumours. Apoptosis was not observed in liver tissue. No apoptosis was observed with the inactive analogue 8-methylxanthenone-4-acetic acid. Positive TUNEL staining of tumour vascular endothelium was evident in one patient in a Phase I clinical trial, from a breast tumour biopsy taken 3 and 24 h after infusion of 5,6-Dimethylxanthenone-4-acetic acid (3.1 mg m−2). Tumour necrosis and the production of tumour tumour necrosis factor were not observed. No apoptotic staining was seen in tumour biopsies taken from two other patients (doses of 3.7 and 4.9 mg m−2). We conclude that 5,6-Dimethylxanthenone-4-acetic acid can induce vascular endothelial cell apoptosis in some murine and human tumours. The action is rapid and appears to be independent of tumour necrosis factor induction.5,6-Dimethylxanthenone-4-acetic acid, synthesised in this laboratory, reduces tumour blood flow, both in mice and in patients on Phase I trial. We used TUNEL (TdT-mediated dUTP nick end labelling) assays to investigate whether apoptosis induction was involved in its antivascular effect. 5,6-Dimethylxanthenone-4-acetic acid induced dose-dependent apoptosis British Journal of Cancer (2002) 86, 1937–1942. doi:10.1038/sj.bjc.6600368www.bjcancer.comCancer Research UK© 2002  DMXAA , a new anticancer agent synthesised in this laboratory  in a volume of 0.01 ml g−1 body weight. DMXAA was dissolved directly in culture medium for in vitro experiments. In the clinical phase trial conducted in Auckland, New Zealand, DMXAA was administered as a 20 min intravenous infusion on a 3-weekly schedule, using a pre-formulated solution of 20 mg ml−1 in 0.1 M phosphate buffer at pH 7.7 (DMXAA was synthesised in this laboratory (3) were implanted subcutaneously in the left flank of anaesthetised  normal C57Bl/6 mice. Tumours were used when they had reached approximately 6 mm in diameter, generally 9–10 days after implantation. Three mice were used for each time point and treatment, and 2–4 cryosections per organ per mouse were immunostained for apoptosis using TUNEL.Athymic or normal C57Bl/6 mice were from the Animal Laboratories, University of Auckland School of Medicine. All mice were maintained under constant temperature and humidity according to institutional ethical guidelines and used between 8–12 weeks of age. All animal experiments have been carried out with ethical committee approval. The ethical guidelines that were followed meet the standards required by the UKCCCR guidelines  consented to biopsies. These samples were taken prior to infusion of DMXAA then at various intervals afterwards and were embedded  and snap-frozen. Representative cryosections were immunostained for apoptosis using TUNEL.2. Sub-confluent cultures (that had been passaged the previous day) were incubated with DMXAA for the appropriate time. The supernatant containing non-adherent cells was removed and centrifuged at 300×g for 10 min. Adherent cells were lifted from the dishes by trypsinisation and collected similarly. Combined adherent and supernatant cells, resuspended in culture medium (0.4 ml), were used to prepared cytospin preparations that were assayed for apoptosis.The HECPP murine endothelial cell line derived from endothelial cells isolated from murine Peyer's patches (In Situ Cell Death Detection Kit (Boehringer Mannheim) according to manufacturer's instructions. Tissue and tumour cryosections or cytospots of cells on poly-L-lysine-treated slides were fixed in 4% paraformaldehyde for 30 min at room temperature, washed with PBS  and then treated with permeabilisation solution (0.1% Triton X-100 in 0.1% sodium citrate) for 2 min on ice. Strand breaks were labelled with fluoresceinated dUTP and visualised following reaction with phosphatase-conjugated antibody to fluorescein and Vector® Black alkaline phosphatase substrate solution . All slides were counter stained using methyl green.Apoptosis was determined using the TUNEL assay for identification of double-stranded DNA breaks using an Cryosections of Colon 38 tumour tissues were also stained for endothelial cells to differentiate between apoptosis of the endothelium and apoptosis of tumour cells. Sections were first processed for apoptosis as described and then immunostained for endothelial cells using a rat anti-mouse CD-31 antibody  . The membranes were UV-crosslinked  and baked (80°C for 30 min). Each membrane was prehybridised  in 7 ml hybridisation mix containing 50% formamide, 0.075 M sodium chloride, 0.05 M sodium dihydrogen phosphate, 5 mM EDTA, 0.001% polyvinyl pyrrolidone, 0.001% bovine serum albumin, 0.001% Ficoll, 0.01 mg ml−1 herring sperm DNA, and 0.5% SDS (sodium dodecyl sulphate). The cDNA to the cytokine gene of interest was labelled with α32P-dCTP (Amersham) using a random priming kit . Excess radioactivity was removed by elution through a G-50 Sephadex column and labelled probe (106 c.p.m. ml−1 hybridisation mix) was then added to the membrane and hybridised for 36 h at 42°C. The blots were washed twice in 2×SSC  with 0.1% SDS for 10 min at 42°C, and finally in 0.2×SSC with 0.1% SDS for 10 min at 65°C. Blots were exposed to X-ray film for 1–3 days at −70°C. After hybridisation with one probe, membranes were stripped (two washes in 300 ml 0.1×SSC with 1% SDS for 15 min at 80°C) and re-hybridised with another probe. The signal intensity was quantitated by laser densitometric scanning. Loading of lanes was determined from the intensity of bands hybridised with the probe for glyceraldehyde-3-phosphate dehydrogenase (GADPH).Total cellular RNA was extracted using RNAzol  according to manufacturer's instructions. RNA (10 μg) was denatured and electrophoresed in 1% agarose-formaldehyde gels as previously described (−1), were double stained for apoptosis and for CD31expression  showed no apoptotic vessels or tumour necrosis (−1)  did not stain with the TUNEL assay  that is achievable in vivo after administration of an effective antitumour dose  of apoptosis of vascular endothelial cells in the Colon 38 tumour. It is not known whether vascular endothelial apoptosis could occur as rapidly in human tumours, since the earliest time point following DMXAA that we were able to examine was 3 h. However, apoptosis occurred more rapidly following DMXAA administration in situ TNF synthesis (Clinical trials of DMXAA have indicated a low response rate to DMXAA (


Accurate and current functional annotation of microarray probes is essential for the analysis and interpretation of the biological processes involved. As gene structures and functional annotation are updated in genome databases, the annotation attached to microarray probes must be updated so that scientists have access to the latest information with which to analyse their data.Eimeria. Using our pipeline, 770 of the probes were assigned at least one entry in either the Ensembl, UniGene or the DFCI gene indices databases.We have designed a pipeline and database for the annotation of microarray probes using publically available databases. The pipeline is based on NCBI BLAST, Perl and MySQL. The pipeline was used to annotate a subset of 791 differentially expressed ArkGenomics chicken probes from an experiment involving chickens infected with the protozoan parasite The pipeline described here provides a simple and robust way of maintaining up-to-date and accurate annotation for microarray probes. The pipeline is designed in such a way as to be flexible and easy to update with new information. The use of microarrays plays an important role in biomedical research, producing large quantities of data on genes that are differentially expressed under various conditions. Although annotation provided with the microarray may be current at the time of manufacture, regular reannotation of the microarray is essential to keep the annotation current. Additionally, probes may be designed from annotation based on incomplete genomes and incorrect or incomplete annotation. This may result in an incomplete coverage of the genome, non-specific probes, incorrect annotation, and orphan probes.ProbeLynx  is a sofThe pipeline has default filters such that only hits that match at greater than 80% identity across at least 20% of the length of the query sequence are counted. These values can be changed depending on requirements; for example, users would choose different values for a cDNA array compared to an oligo array. With these default values, 770 probes had at least one matching hit in at least one of the Ensembl, UniGene or DFCI gene indices databases.Applying the selection criteria to the data presented here resulted in 750 probes having at least one matching hit in at least one of the Ensembl , UniGeneUsing the Ensembl database, annotation could be provided for 472 probes (60%). Of those, 438 matched a single Ensembl gene id and 34 probes matched multiple genes. A total of 426 probes had perfect matches. Of these, 396 were unique hits. Gene descriptions were provided for 405 probes and 198 probes were matched to at least one Gene Ontology [Using the DFCI gene indices, annotation was provided for 683 probes (86%). Of these, 249 matched a single gene index, 434 matched multiple indices, and 548 probes had perfect matches, 195 of which had single unique hits. Using the DFCI gene indices annotation, a gene description was provided for 466 probes and 66 probes were matched to at least one GO term .Of the 791 probes, 715 (90%) could be assigned to at least one UniGene cluster, of which 593 were assigned uniquely (and therefore 122 were assigned to multiple clusters). Perfect matches were seen in 560 cases, of which 478 were unique. All 715 of the annotated probes had a cluster title (gene description).When linking microarray probes to genome databases, we are attempting to do two things. Firstly, we are attempting to define just how many genes might be hybridising to each spot and contributing to the signal intensity. Secondly, we are attempting to inform scientists about gene function.Ideally there should be a one-to-one relationship between probe and gene. However, this is clearly not the case. Using the selection criteria, the best results come from UniGene, where 75% of probes have a single, contributory gene; the worst results are from DFCI gene indices, where the figure is 31%. Probes with more than one hit may be due to shared domains, overlapping genes, misannotation, misassembly, low complexity regions, and/or repeat regions.There are several reasons why probes may have no hits. The microarray used in this study was designed in 2005 using the first draft of the chicken genome, Ensembl version 30, and annotated with Ensembl version 42. Since then there have been 20 subsequent versions of the Ensembl database and a second draft of the chicken genome. Regular reannotation of the probes using the information provided with new genebuilds and Ensembl releases allows us to maintain up-to-date information. In addition, only the core Ensembl gene set was searched; had we searched against the genome itself, or against the EST gene set, the number of unannotated probes would be reduced. It is not surprising that the number of unannotated probes is lower in the two EST databases. However, even with UniGene, the best in terms of probe coverage, one in ten probes did not have a hit above the threshold. This may mean that the sequence that the probe was originally designed to is no longer publicly available (or never was) or that it did not meet the quality criteria applied before the database was built.In terms of functional annotation, all three databases provided a functional description for over half of the probes. UniGene again performed the best, although no attempt has been made to judge the quality of the description. Disappointingly, a maximum of 25% of probes were assigned GO terms.Future improvements in the assembly of the chicken genome and annotation should help to increase the level of annotation. The IMAD pipeline could be improved by allowing searches against the genome assembly, and against further databases such as the Ensembl EST genes, KEGG , and RefWe have created a pipeline that can be used to maintain the annotation of microarray probes using publicly available databases. The analysis of a set of differentially expressed probes revealed problems with annotation that may be due to a probe design based on incomplete annotation of the chicken genome. As improvements in the annotation of the chicken genome are made, improvements in the design of chicken microarrays are sure to follow.IMAD consists of a flexible relational database in MySQL, designed to store the hits of any set of sequences against any number of BLAST  databaseThe probe set was searched against multiple databases using NCBI BLAST, followed by parsing of the BLAST results. Where a single HSP exists between the query and hit, filters are applied and statistics are calculated and stored in the database. Where there are multiple HSPs, any overlap with respect to the query and the hit is removed. Statistics are then applied across all HSPs, filters applied and then stored in the database. Results (top hit for each probe for each database) in spreadsheet format are extracted using the API.Eimeria infected chickens [The microarray used in this study was the Arkgenomics chicken 20 K oligo microarray, consisting 20,460 probes designed against a unique set of chicken transcripts in 2005, primarily 70 mer oligos . A subsechickens .Ensembl chicken version 50, UniGene chicken build 39 and DFCI chicken gene indices version 11 were used. Gene Ontology terms were obtained through Ensembl BioMart . These tCutoff values for positive hits were any target with a contiguous matching stretch greater than 20 bases and an overall percentage identity greater than 80%. A perfect match is defined where there is a 100% match over the entire length of the oligo with the target sequence.The authors declare that they have no competing interests.MW and DP co-wrote IMAD.


Elucidation of the basic genetic changes of human hepatocellular carcinoma is important for the understanding and treatment of this cancer. We used microsatellite polymorphism markers to study 30 cases of hepatocellular carcinoma (34 tumours) on all human chromosomes. DNA from 34 pairs of hepatocellular carcinomas and corresponding non-tumour parts was prepared. Loss of heterozygosity (LOH) and microsatellite instability on 23 chromosomes were investigated by 231 sets of microsatellite markers. More than 20% LOH was shown for loci on 16q (47.1%), 13q (32.4%), 17p (32.4%), 5q (26.5%), 11p (23.5%) and 9p (20.6%). The commonly affected regions were mapped to 16q12.1, 16q12.2, 16q24, 13q12.1–32, 17p13, 5q32, 5q34, 5q3, 11p15, 11q23–24 and 9p21. Hepatitis B virus carriers had a significantly higher frequency of LOH on chromosomes 5q, 11p and 16q. Furthermore, larger tumour size tended to have higher frequency of LOH at D16S409 locus (16q12.1). Microsatellte instability was only found in 12 of 231 markers and the frequency is very low. These data suggest that the chromosomes 16q, 13q, 17p, 5q, 11p and 9p might participate in hepatocarcinogenesis. However, microsatellite instability might play little role in the development of this cancer in Taiwan. © 1999 Cancer Research Campaign


The aim of the study was to establish a model of tumour microcirculation in vivo using the murine renal cell carcinoma cell line (RENCA) implanted into the mouse cremaster muscle, and subsequently to investigate the trafficking of syngeneic lymphocyte subpopulations into both the RENCA tumour and the surrounding normal cremaster muscle microcirculation. We have demonstrated that RENCA tumour cells, at a dose of 1.5 x 10(5) per 30 microl injected into the cremaster muscle, reproducibly produced a vascularized tumour suitable for in vivo microscopy at 10-14 days. Injection of fluorescently labelled effector cells (1 x 10(6)) including naive splenocytes, T-cell enriched populations and ex vivo interleukin 2 (IL-2)-activated splenocytes all migrated to and flowed through both the tumour and the normal microcirculation, with negligible adhesion. However, we observed the selective recruitment, localization and arrest of IL-2-activated splenocytes (P < 0.05) into the tumour microcirculation, and the subsequent extravasation of cells into the tumour intestitium in some instances. This did not occur with the other effector cells. We also observed the absence of leucocyte rolling in the tumour microcirculation, suggesting an impairment in adhesion molecule expression on the tumour endothelium. We have therefore established the potential of this model for defining further effector cell-tumour-endothelium interactions.


The results showed a transitory increase in thymidine labelling index 12 hours after drug injection accompanied by a low mitotic index which persisted for a further 12 hours. These observations are compared with, and confirm, a previous study involving analysis of smear preparations from tumour cell suspensions. The changes in the proportion of the tumour occupied by erythrocytes and necrotic elements are described. The results of the morphometric analysis have shown the changes occurring in the outer, intermediate and inner zones of the tumour. In all control tumours, and tumours up to 2 days after cyclophosphamide treatment, necrosis was predominant in the inner zone and tumour cell proliferation was most marked in the outer zone. In contrast, 3 days after cyclophosphamide treatment proliferation was predominant in the inner zone and the amount of necrosis was approximately the same in each zone.The response of a transplantable rat fibrosarcoma, RIB


Single cell suspensions of two allogeneic tumours (W-256 and Y-P388) injected intravenously produced macrocolonies in the lungs of rats. Colony forming efficiency  was low in 6-week or older rats but was markedly increased by 1000-1500 rad local thoracic irradiation (LTI) given 7-14 days before the tumour cell injection, or by antilymphocytic serum (ALS) but not by sublethal whole body irradiation (WBI). Similarly, LTI increased the incidence of pulmonary metastases produced by a solid tumour growing in the leg muscle. Stimulation of CFE by LTI was a strictly local phenomenon and not due to effects of irradiation on thymus, spleen or other tissues of the rat. LTI failed to increase CFE in immunized rats. It is concluded that (1) LTI stimulates clonogenic growth of tumour cells arrested in the lungs, by causing inflammatory reactions accompanied by regenerative cellular proliferation of lung tissue, which increases the “plating” efficiency of tumour cells, (2) the increase in CFE in lungs is not due to suppression of immunity to tumour growth by LTI.