New prognostic factors and calculators for outcome prediction in patients with recurrent glioblastoma: A pooled analysis of EORTC Brain Tumour Group phase I and II clinical trials.

BACKGROUND: Prognostic models have been developed to predict survival of patients with newly diagnosed glioblastoma (GBM). To improve predictions, models should be updated with information at the recurrence. We performed a pooled analysis of European Organization for Research and Treatment of Cancer (EORTC) trials on recurrent glioblastoma to validate existing clinical prognostic factors, identify new markers, and derive new predictions for overall survival (OS) and progression free survival (PFS).¦METHODS: Data from 300 patients with recurrent GBM recruited in eight phase I or II trials conducted by the EORTC Brain Tumour Group were used to evaluate patient's age, sex, World Health Organisation (WHO) performance status (PS), presence of neurological deficits, disease history, use of steroids or anti-epileptics and disease characteristics to predict PFS and OS. Prognostic calculators were developed in patients initially treated by chemoradiation with temozolomide.¦RESULTS: Poor PS and more than one target lesion had a significant negative prognostic impact for both PFS and OS. Patients with large tumours measured by the maximum diameter of the largest lesion (⩾42mm) and treated with steroids at baseline had shorter OS. Tumours with predominant frontal location had better survival. Age and sex did not show independent prognostic values for PFS or OS.¦CONCLUSIONS: This analysis confirms performance status but not age as a major prognostic factor for PFS and OS in recurrent GBM. Patients with multiple and large lesions have an increased risk of death. With these data prognostic calculators with confidence intervals for both medians and fixed time probabilities of survival were derived.

Awakening dormant haematopoietic stem cells.

Haematopoietic stem cells (HSCs) in mouse bone marrow are located in specialized niches as single cells. During homeostasis, signals from this environment keep some HSCs dormant, which preserves long-term self-renewal potential, while other HSCs actively self renew to maintain haematopoiesis. In response to haematopoietic stress, dormant HSCs become activated and rapidly replenish the haematopoietic system. Interestingly, three factors - granulocyte colony-stimulating factor, interferon-alpha and arsenic trioxide - have been shown to efficiently activate dormant stem cells and thereby could break their resistance to anti-proliferative chemotherapeutics. Thus, we propose that two-step strategies could target resistant leukaemic stem cells by priming tumours with activators of dormancy followed by chemotherapy or targeted therapies.

Skull base chordomas: presentation of six cases and review of the literature.

Chordomas are uncommon tumours occuring from remnants of the notochord. They are mainly localized in the sacrococcygium, the spine and the skull base. We focused our study on skull base chordomas and reviewed 531 published cases and 6 cases diagnosed in our institution. The six cases diagnosed at our hospital are described with a special mention of a unique case presenting with meningitis. Clinical features, radiology, histopathology and survival with different treatments are reviewed. Skull base chordomas can occur at any age and are slightly more frequent in males. Patients complain most often of diplopia and headaches. Abducens nerve palsy is the commonest neurological finding. Radiological examination should include both CT-scan which shows extensive osteolysis, calcifications and contrast enhancement and MRI for topographical definition. Metastases rarely occur. Treatment remains controversial but combined surgery and radiotherapy is the best treatment with a 5 year survival rate of 75%.

Workers exposed to wood dust have an increased micronucleus frequency in nasal and buccal cells: results from a pilot study

Wood dust is recognised as a human carcinogen, based on the strong association of wood dust exposure and the elevated risk of malignant tumours of the nasal cavity and paranasal sinuses [sino-nasal cancer (SNC)]. The study aimed to assess genetic damage in workers exposed to wood dust using biomarkers in both buccal and nasal cells that reflect genome instability events, cellular proliferation and cell death frequencies. Nasal and buccal epithelial cells were collected from 31 parquet layers, installers, carpenters and furniture workers (exposed group) and 19 non-exposed workers located in Switzerland. Micronucleus (MN) frequencies were scored in nasal and buccal cells collected among woodworkers. Other nuclear anomalies in buccal cells were measured through the use of the buccal micronucleus cytome assay. MN frequencies in nasal and buccal cells were significantly higher in the exposed group compared to the non-exposed group; odds ratio for nasal cells 3.1 [95% confidence interval (CI) 1.8-5.1] and buccal cells 1.8 (95% CI 1.3-2.4). The exposed group had higher frequencies of cells with nuclear buds, karyorrhectic, pyknotic, karyolytic cells and a decrease in the frequency of basal, binucleated and condensed cells compared to the non-exposed group. Our study confirms that woodworkers have an elevated risk for chromosomal instability in cells of the aerodigestive tract. The MN assay in nasal cells may become a relevant biomonitoring tool in the future for early detection of SNC risk. Future studies should seek to standardise the protocol for MN frequency in nasal cells similar to that for MN in buccal cells.

The Wnt receptor FZD1 mediates chemoresistance in neuroblastoma through activation of the Wnt/beta-catenin pathway.

The development of chemoresistance represents a major obstacle in the successful treatment of cancers such as neuroblastoma (NB), a particularly aggressive childhood solid tumour. The mechanisms underlying the chemoresistant phenotype in NB were addressed by gene expression profiling of two doxorubicin (DoxR)-resistant vs sensitive parental cell lines. Not surprisingly, the MDR1 gene was included in the identified upregulated genes, although the highest overexpressed transcript in both cell lines was the frizzled-1 Wnt receptor (FZD1) gene, an essential component of the Wnt/beta-catenin pathway. FZD1 upregulation in resistant variants was shown to mediate sustained activation of the Wnt/beta-catenin pathway as revealed by nuclear beta-catenin translocation and target genes transactivation. Interestingly, specific micro-adapted short hairpin RNA (shRNAmir)-mediated FZD1 silencing induced parallel strong decrease in the expression of MDR1, another beta-catenin target gene, revealing a complex, Wnt/beta-catenin-mediated implication of FZD1 in chemoresistance. The significant restoration of drug sensitivity in FZD1-silenced cells confirmed the FZD1-associated chemoresistance. RNA samples from 21 patient tumours (diagnosis and postchemotherapy), showed a highly significant FZD1 and/or MDR1 overexpression after treatment, underlining a role for FZD1-mediated Wnt/beta-catenin pathway in clinical chemoresistance. Our data represent the first implication of the Wnt/beta-catenin pathway in NB chemoresistance and identify potential new targets to treat aggressive and resistant NB.

Carcinoembryonic antigen expression, antibody localisation and immunophotodetection of human colon cancer liver metastases in nude mice: a model for radioimmunotherapy.

Colorectal cancer frequently disseminates through the portal vein into the liver. In this study, outbred Swiss nude mice were adapted to facilitate the induction of liver metastases by a pre-grafting treatment with 6 Gy total body irradiation and i.v. injection of anti-asialo GM1 antibody. One day later, cultured LS 174T human colon cancer cells were injected into the surgically exposed spleen, which was resected 3 min later. In 48 of 65 mice, a few to several hundred liver metastases were macroscopically observed at dissection 3 to 4 weeks after transplantation. Ten of 10 mice, followed-up for survival, died with multiple large confluent liver metastases. By reducing the radiation dose to 4 or 0 Gy, or omitting the anti-asialo GM1 antibody injection, only 60%, 37% or 50% of mice, respectively, had visible metastases 3 weeks after transplantation. Carcinoembryonic antigen (CEA) measured in tumour extracts was in the mean 25.6 micrograms/g in liver metastases compared with 9.2 micrograms/g in s.c. tumours. Uptake of radiolabelled anti-CEA monoclonal antibody (MAb) in the metastases 12, 24 and 48 hr after injection gave a mean value of 39% of the injected dose per gram of tissue (ID/g). In comparison, MAb uptake in s.c. and intrasplenic tumours or lung metastases gave a mean percentage ID/g of 20, 18 and 15, respectively. Laser-induced fluorescence after injection of indocyanin-MAb conjugate allowed direct visual detection of small liver metastases, including some that were not visible under normal light. Preliminary results showed that mice, pre-treated with 4 Gy irradiation and the anti-asialo GM1 injection, were tolerant to radioimmunotherapy with a total dose of 500 muCi 131I labeled anti-CEA intact MAbs given in 3 injections.

Intensified chemotherapy with ACVBP plus rituximab versus standard CHOP plus rituximab for the treatment of diffuse large B-cell lymphoma (LNH03-2B): an open-label randomised phase 3 trial.

BACKGROUND: The outcome of diffuse large B-cell lymphoma has been substantially improved by the addition of the anti-CD20 monoclonal antibody rituximab to chemotherapy regimens. We aimed to assess, in patients aged 18-59 years, the potential survival benefit provided by a dose-intensive immunochemotherapy regimen plus rituximab compared with standard treatment plus rituximab. METHODS: We did an open-label randomised trial comparing dose-intensive rituximab, doxorubicin, cyclophosphamide, vindesine, bleomycin, and prednisone (R-ACVBP) with subsequent consolidation versus standard rituximab, doxorubicin, cyclophosphamide, vincristine, and prednisone (R-CHOP). Random assignment was done with a computer-assisted randomisation-allocation sequence with a block size of four. Patients were aged 18-59 years with untreated diffuse large B-cell lymphoma and an age-adjusted international prognostic index equal to 1. Our primary endpoint was event-free survival. Our analyses of efficacy and safety were of the intention-to-treat population. This study is registered with ClinicalTrials.gov, number NCT00140595. FINDINGS: One patient withdrew consent before treatment and 54 did not complete treatment. After a median follow-up of 44 months, our 3-year estimate of event-free survival was 81% (95% CI 75-86) in the R-ACVBP group and 67% (59-73) in the R-CHOP group (hazard ratio [HR] 0·56, 95% CI 0·38-0·83; p=0·0035). 3-year estimates of progression-free survival (87% [95% CI, 81-91] vs 73% [66-79]; HR 0·48 [0·30-0·76]; p=0·0015) and overall survival (92% [87-95] vs 84% [77-89]; HR 0·44 [0·28-0·81]; p=0·0071) were also increased in the R-ACVBP group. 82 (42%) of 196 patients in the R-ACVBP group experienced a serious adverse event compared with 28 (15%) of 183 in the R-CHOP group. Grade 3-4 haematological toxic effects were more common in the R-ACVBP group, with a higher proportion of patients experiencing a febrile neutropenic episode (38% [75 of 196] vs 9% [16 of 183]). INTERPRETATION: Compared with standard R-CHOP, intensified immunochemotherapy with R-ACVBP significantly improves survival of patients aged 18-59 years with diffuse large B-cell lymphoma with low-intermediate risk according to the International Prognostic Index. Haematological toxic effects of the intensive regimen were raised but manageable. FUNDING: Groupe d'Etudes des Lymphomes de l'Adulte and Amgen.

Mutations in the EXT1 and EXT2 genes in Spanish patients with multiple osteochondromas

Multiple osteochondromas is an autosomal dominant skeletal disorder characterized by the formation of multiple cartilage-capped tumours. Two causal genes have been identified, EXT1 and EXT2, which account for 65% and 30% of cases, respectively. We have undertaken a mutation analysis of the EXT1 and EXT2 genes in 39 unrelated Spanish patients, most of them with moderate phenotype, and looked for genotype-phenotype correlations. We found the mutant allele in 37 patients, 29 in EXT1 and 8 in EXT2. Five of the EXT1 mutations were deletions identified by MLPA. Two cases of mosaicism were documented. We detected a lower number of exostoses in patients with missense mutation versus other kinds of mutations. In conclusion, we found a mutation in EXT1 or in EXT2 in 95% of the Spanish patients. Eighteen of the mutations were novel.

Mutations in the EXT1 and EXT2 genes in Spanish patients with multiple osteochondromas

Multiple osteochondromas is an autosomal dominant skeletal disorder characterized by the formation of multiple cartilage-capped tumours. Two causal genes have been identified, EXT1 and EXT2, which account for 65% and 30% of cases, respectively. We have undertaken a mutation analysis of the EXT1 and EXT2 genes in 39 unrelated Spanish patients, most of them with moderate phenotype, and looked for genotype-phenotype correlations. We found the mutant allele in 37 patients, 29 in EXT1 and 8 in EXT2. Five of the EXT1 mutations were deletions identified by MLPA. Two cases of mosaicism were documented. We detected a lower number of exostoses in patients with missense mutation versus other kinds of mutations. In conclusion, we found a mutation in EXT1 or in EXT2 in 95% of the Spanish patients. Eighteen of the mutations were novel.

T cell differentiation in chronic infection and cancer: functional adaptation or exhaustion?

Chronic viral infections and malignant tumours induce T cells that have a reduced ability to secrete effector cytokines and have upregulated expression of the inhibitory receptor PD1 (programmed cell death protein 1). These features have so far been considered to mark terminally differentiated 'exhausted' T cells. However, several recent clinical and experimental observations indicate that phenotypically exhausted T cells can still mediate a crucial level of pathogen or tumour control. In this Opinion article, we propose that the exhausted phenotype results from a differentiation process in which T cells stably adjust their effector capacity to the needs of chronic infection. We argue that this phenotype is optimized to cause minimal tissue damage while still mediating a critical level of pathogen control. In contrast to the presently held view of functional exhaustion, this new concept better reflects the pathophysiology and clinical manifestations of persisting infections, and it provides a rationale for emerging therapies that enhance T cell activity in chronic infection and cancer by blocking inhibitory receptors.

Successful migration of three tracers without identification of sentinel nodes during intraoperative lymphatic mapping for non-small cell lung cancer.

Prospective comparative evaluation of patent V blue, fluorescein and (99m)TC-nanocolloids for intraoperative sentinel lymph node (SLN) mapping during surgery for non-small cell lung cancer (NSCLC). Ten patients with peripherally localised clinical stage I NSCLC underwent thoracotomy and peritumoral subpleural injection of 2 ml of patent V blue dye, 1 ml of 10% fluorescein and 1ml of (99m)Tc-nanocolloids (0.4 mCi). The migration and spatial distribution pattern of the tracers was assessed by direct visualisation (patent V blue), visualisation of fluorescence signalling by a lamp of Wood (fluorescein) and radioactivity counting with a hand held gamma-probe ((99m)Tc-nanocolloids). Lymph nodes at interlobar (ATS 11), hilar (ATS 10) and mediastinal (right ATS 2,4,7; left ATS 5,6,7) levels were systematically assessed every 10 min up to 60 min after injection, followed by lobectomy and formal lymph node dissection. Successful migration from the peritumoral area to the mediastinum was observed for all three tracers up to 60 min after injection. The interlobar lympho-fatty tissue (station ATS 11) revealed an early and preferential accumulation of all three tracers for all tumours assessed and irrespective of the tumour localisation. However, no preferential accumulation in one or two distinct lymph nodes was observed up to 60 min after injection for all three tracers assessed. Intraoperative SLN mapping revealed successful migration of the tracers from the site of peritumoral injection to the mediastinum, but in a diffuse pattern without preferential accumulation in sentinel lymph nodes.

Combining hypoxic methods for peak performance.

New methods and devices for pursuing performance enhancement through altitude training were developed in Scandinavia and the USA in the early 1990s. At present, several forms of hypoxic training and/or altitude exposure exist: traditional 'live high-train high' (LHTH), contemporary 'live high-train low' (LHTL), intermittent hypoxic exposure during rest (IHE) and intermittent hypoxic exposure during continuous session (IHT). Although substantial differences exist between these methods of hypoxic training and/or exposure, all have the same goal: to induce an improvement in athletic performance at sea level. They are also used for preparation for competition at altitude and/or for the acclimatization of mountaineers. The underlying mechanisms behind the effects of hypoxic training are widely debated. Although the popular view is that altitude training may lead to an increase in haematological capacity, this may not be the main, or the only, factor involved in the improvement of performance. Other central (such as ventilatory, haemodynamic or neural adaptation) or peripheral (such as muscle buffering capacity or economy) factors play an important role. LHTL was shown to be an efficient method. The optimal altitude for living high has been defined as being 2200-2500 m to provide an optimal erythropoietic effect and up to 3100 m for non-haematological parameters. The optimal duration at altitude appears to be 4 weeks for inducing accelerated erythropoiesis whereas <3 weeks (i.e. 18 days) are long enough for beneficial changes in economy, muscle buffering capacity, the hypoxic ventilatory response or Na(+)/K(+)-ATPase activity. One critical point is the daily dose of altitude. A natural altitude of 2500 m for 20-22 h/day (in fact, travelling down to the valley only for training) appears sufficient to increase erythropoiesis and improve sea-level performance. 'Longer is better' as regards haematological changes since additional benefits have been shown as hypoxic exposure increases beyond 16 h/day. The minimum daily dose for stimulating erythropoiesis seems to be 12 h/day. For non-haematological changes, the implementation of a much shorter duration of exposure seems possible. Athletes could take advantage of IHT, which seems more beneficial than IHE in performance enhancement. The intensity of hypoxic exercise might play a role on adaptations at the molecular level in skeletal muscle tissue. There is clear evidence that intense exercise at high altitude stimulates to a greater extent muscle adaptations for both aerobic and anaerobic exercises and limits the decrease in power. So although IHT induces no increase in VO(2max) due to the low 'altitude dose', improvement in athletic performance is likely to happen with high-intensity exercise (i.e. above the ventilatory threshold) due to an increase in mitochondrial efficiency and pH/lactate regulation. We propose a new combination of hypoxic method (which we suggest naming Living High-Training Low and High, interspersed; LHTLHi) combining LHTL (five nights at 3000 m and two nights at sea level) with training at sea level except for a few (2.3 per week) IHT sessions of supra-threshold training. This review also provides a rationale on how to combine the different hypoxic methods and suggests advances in both their implementation and their periodization during the yearly training programme of athletes competing in endurance, glycolytic or intermittent sports.

Development of Craniofacial Bone Defect Reconstruction Implant Based on Fibre-Reinforced Composite with Photopolymerisable Resin Systems. Experimental studies in vitro and in vivo.

Reconstruction of defects in the craniomaxillofacial (CMF) area has mainly been based on bone grafts or metallic fixing plates and screws. Particularly in the case of large calvarial and/or craniofacial defects caused by trauma, tumours or congenital malformations, there is a need for reliable reconstruction biomaterials, because bone grafts or metallic fixing systems do not completely fulfill the criteria for the best possible reconstruction methods in these complicated cases. In this series of studies, the usability of fibre-reinforced composite (FRC) was studied as a biostable, nonmetallic alternative material for reconstructing artificially created bone defects in frontal and calvarial areas of rabbits. The experimental part of this work describes the different stages of the product development process from the first in vitro tests with resin-impregnated fibrereinforced composites to the in vivo animal studies, in which this FRC was tested as an implant material for reconstructing different size bone defects in rabbit frontal and calvarial areas. In the first in vitro study, the FRC was polymerised in contact with bone or blood in the laboratory. The polymerised FRC samples were then incubated in water, which was analysed for residual monomer content by using high performance liquid chromatography (HPLC). It was found that this in vitro polymerisation in contact with bone and blood did not markedly increase the residual monomer leaching from the FRC. In the second in vitro study, different adhesive systems were tested in fixing the implant to bone surface. This was done to find an alternative implant fixing system to screws and pins. On the basis of this study, it was found that the surface of the calvarial bone needed both mechanical and chemical treatments before the resinimpregnated FRC could be properly fixed onto it. In three animal studies performed with rabbit frontal bone defects and critical size calvarial bone defect models, biological responses to the FRC implants were evaluated. On the basis of theseevaluations, it can be concluded that the FRC, based on E-glass (electrical glass) fibres forming a porous fibre veil enables the ingrowth of connective tissues to the inner structures of the material, as well as the bone formation and mineralization inside the fibre veil. Bone formation could be enhanced by using bioactive glass granules fixed to the FRC implants. FRC-implanted bone defects healed partly; no total healing of defects was achieved. Biological responses during the follow-up time, at a maximum of 12 weeks, to resin-impregnated composite implant seemed to depend on the polymerization time of the resin matrix of the FRC. Both of the studied resin systems used in the FRC were photopolymerised and the heat-induced postpolymerisation was used additionally.

Effects of preset sequential administrations of sunitinib and everolimus on tumour differentiation in Caki-1 renal cell carcinoma.

BACKGROUND: Sunitinib (VEGFR/PDGFR inhibitor) and everolimus (mTOR inhibitor) are both approved for advanced renal cell carcinoma (RCC) as first-line and second-line therapy, respectively. In the clinics, sunitinib treatment is limited by the emergence of acquired resistance, leading to a switch to second-line treatment at progression, often based on everolimus. No data have been yet generated on programmed alternating sequential strategies combining alternative use of sunitinib and everolimus before progression. Such strategy is expected to delay the emergence of acquired resistance and improve tumour control. The aim of our study was to assess the changes in tumours induced by three different sequences administration of sunitinib and everolimus. METHODS: In human Caki-1 RCC xenograft model, sunitinib was alternated with everolimus every week, every 2 weeks, or every 3 weeks. Effects on necrosis, hypoxia, angiogenesis, and EMT status were assessed by immunohisochemistry and immunofluorescence. RESULTS: Sunitinib and everolimus programmed sequential regimens before progression yielded longer median time to tumour progression than sunitinib and everolimus monotherapies. In each group of treatment, tumour growth control was associated with inhibition of mTOR pathway and changes from a mesenchymal towards an epithelial phenotype, with a decrease in vimentin and an increase in E-cadherin expression. The sequential combinations of these two agents in a RCC mouse clinical trial induced antiangiogenic effects, leading to tumour necrosis. CONCLUSIONS: In summary, our study showed that alternate sequence of sunitinib and everolimus mitigated the development of mesenchymal phenotype compared with sunitinib as single agent.

Direct comparison of a radioiodinated intact chimeric anti-CEA MAb with its F(ab')2 fragment in nude mice bearing different human colon cancer xenografts.

Tumour localisation and tumour to normal tissue ratios of a chimeric anti-carcinoembryonic antigen (CEA) monoclonal antibody (MAb), in intact form and as an F(ab')2 fragment labelled with 125I and 131I, were compared in groups of nude mice bearing four different colon cancer xenografts, T380, Co112 or LoVo, of human origin, or a rat colon cancer transfected with human CEA cDNA, called '3G7'. For each tumour, three to four mice per time point were analysed 6, 12, 24, 48 and 96 h after MAb injection. In the different tumours, maximal localisation of intact MAb was obtained at 24 to 48 h, and of F(ab')2 fragment 12 to 24 h after injection. Among the different tumours, localisation was highest with colon cancer T380, with 64% of the injected dose per gram (% ID/g) for the intact MAb and 57% for its F(ab')2 fragment, while in the three other tumours, maximal localisation ranged from 14 to 22% ID g-1 for the intact MAb and was about 11% for the F(ab')2. Tumour to normal tissue ratios of intact MAb increased rapidly until 24 h after injection and remained stable or showed only a minor increase thereafter. In contrast, for the F(ab')2 fragment, the tumour to normal tissue ratios increased steadily up to 4 days after injection reaching markedly higher values than those obtained with intact MAb. For the four different xenografts, tumour to blood ratios of F(ab')2 were about 2, 3 and 5 to 16 times higher than those of intact antibodies at 12, 24 and 96 h after injection, respectively.