Breast and ovarian cancer predisposition due to de novo BRCA1 and BRCA2 mutations.

BRCA1 and BRCA2 are the two major genes predisposing to breast and ovarian cancer. Whereas high de novo mutation rates have been demonstrated for several genes, only 11 cases of de novo BRCA1/2 mutations have been reported to date and the BRCA1/2 de novo mutation rate remains unknown. The present study was designed to fill this gap based on a series of 12 805 consecutive unrelated patients diagnosed with breast and/or ovarian cancer who met the inclusion criteria for BRCA1/2 gene analysis according to French guidelines. BRCA1/2 mutations were detected in 1527 (12%) patients, and three BRCA1 mutations and one BRCA2 mutation were de novo. The BRCA1/2 de novo mutation rate was estimated to be 0.3% (0.1%; 0.7%). Although rare, it may be useful to take the possibility of de novo BRCA1/2 mutation into account in genetic counseling of relatives and to improve the understanding of complex family histories of breast and ovarian cancers.

Productivity, water-use efficiency and tolerance to moderate water deficit correlate in 33 poplar genotypes from a Populus deltoides x Populus trichocarpa F1 progeny.

Genotypic variability for productivity, water-use efficiency and leaf traits in 33 genotypes selected from an F1 progeny of Populus deltoides Bartr. ex Marsh x Populus trichocarpa L. was explored under optimal and moderate water-deficit conditions. Saplings of the 33 genotypes were grown in a two-plot open field at INRA Orléans (France) and coppiced every year. A moderate water deficit was induced during two successive years on one plot by withholding irrigation, while the second one remained irrigated (control). Stem biomass and leaf structure (e.g., specific leaf area and leaf area) were measured in 2004 and 2005 and functional leaf traits (e.g., carbon isotope discrimination, Delta) were measured only in 2004. Tolerance to water deficit was estimated at genotype level as the ability to limit losses in biomass production in water deficit versus control trees. Stem biomass, leaf structure and Delta displayed a significant genotypic variability whatever the irrigation regime. For all traits, genotype ranks remained stable across years for similar irrigation conditions. Carbon isotope discrimination scaled negatively with productivity and leaf nitrogen content in controls. The most productive genotypes were the least tolerant to moderate water deficit. No relationship was evidenced between Delta and the level of tolerance to water deficit. The relationships between traits evidenced in this collection of P. deltoides x P. trichocarpa F1 genotypes contrast with the ones that were previously detected in a collection of P. deltoides x Populus nigra L. cultivars tested in the same field trial.

The carbon isotope composition of CO2 respired by trunks: comparison of four sampling methods.

The (13)C natural abundance of CO(2) respired by plants has been used in the laboratory to examine the discrimination processes that occur during respiration. Currently, field measurements are being expanded to interpret the respiration delta(13)C signature measured at ecosystem and global levels. In this context, forests are particularly important to consider as they represent 80% of the continental biomass. The objective of this investigation was to compare four methods of sampling the CO(2) respired by trunks for the determination of its carbon isotope composition: three in situ methods using chambers placed on the trunk, and one destructive method using cores of woody tissues. The in situ methods were based either on a Keeling plot approach applied at the tissue level or on an initial flush of the chamber with nitrogen or with CO(2)-free air. In parallel, we investigated the possibility of an apparent discrimination during tissue respiration by comparing the delta(13)C signature of the respired CO(2) and that of the organic matter. The study was performed on six tree species widely distributed in temperate and mediterranean areas. The four methods were not significantly different when overall means were considered. However, considering the individual data, the Keeling plot approach and the nitrogen flush methods gave fairly homogeneous results, whereas the CO(2)-free air method produced more variable results. The core method was not correlated with any of the chamber methods. Regardless of the methodology, the respired CO(2) generally was enriched in (13)C relative to the total organic matter. This apparent enrichment during respiration was variable, reaching as much as 3-5 per thousand. This study showed that, on the whole, the different sampling techniques gave similar results, but one should be aware of the variability associated with each method.

Distribution of above-ground and below-ground carbohydrate reserves in adult trees of two contrasting broad-leaved species (Quercus petraea and Fagus sylvatica).

• The exhaustive distribution of total carbohydrate reserves was investigated in oak and beech trees that were approx. 40 yr old and felled at two dates (October 1999 and June 2000) to estimate variations in reserve amounts at the tree level. • The total nonstructural carbohydrate (TNC) content was highest in the twigs and coarse roots, reaching 10 g 100 g-1 dry matter and 12 g 100 g-1 dry matter for beech and oak twigs, and 13 g 100 g-1 dry matter and 16 g 100 g-1 dry matter for beech and oak roots, respectively. Similar distribution in tree carbohydrates was observed for both species and date, but with contrasting starch/sugar sharing. • Scaling-up to reserve amounts at tree level was performed with extensive organ biomass measurements. Based on the respective biomass of the organs, stem and roots contained the highest quantity of reserves. Between October (before leaf fall) and June (after bud-burst and leaf area index expansion) oaks used double the reserves of beeches. • These differences in the allocation of carbohydrate reserves could arise from differential needs for spring growth and winter maintenance respiration between the two species.

Contrasting distribution and seasonal dynamics of carbohydrate reserves in stem wood of adult ring-porous sessile oak and diffuse-porous beech trees.

We tested the hypothesis that broad-leaved forest species with contrasting wood anatomy and hydraulic system (ring-porous versus diffuse-porous) also differ in distribution and seasonal dynamics of carbohydrate reserves in stem wood. Total nonstructural carbohydrate (TNC) reserves (starch and sugars) were measured enzymatically in the 10 youngest stem xylem rings of adult oak (Quercus petraea (Matt.) Liebl.) and beech (Fagus sylvatica L.) trees during an annual cycle. Radial distribution of carbohydrates was investigated according to ring age. On all dates, oak trees had twofold higher TNC concentration than beech trees (41 versus 23 mg g(DM)(-1)), with starch accounting for the high TNC concentration in oak. Seasonal dynamics of TNC concentration were significantly (P < 0.05) more pronounced in oak (20-64 mg TNC g(DM)(-1)) than in beech (17-34 mg TNC g(DM)(-1)). A marked decrease in TNC concentration was observed in oak trees during bud burst and early wood growth, whereas seasonal fluctuations in TNC concentrations in beech trees were small. The radial distribution of TNC based on ring age differed between species: TNC was restricted to the sapwood rings in oak, whereas in beech, it was distributed throughout the wood from the outermost sapwood ring to the pith. Although the high TNC concentrations in the outermost rings accounted for most of the observed seasonal pattern, all of the 10 youngest xylem rings analyzed participated in the seasonal dynamics of TNC in beech trees. The innermost sapwood rings of oak trees had low TNC concentrations. Stem growth and accumulation of carbon reserves occurred concomitantly during the first part of the season, when there was no soil water deficit. When soil water content was depleted, stem growth ceased in both species, whereas TNC accumulation was negligibly affected and continued until leaf fall. The contrasting dynamics and distribution of carbohydrate reserves in oak and beech are discussed with reference to differences in phenology, early spring growth and hydraulic properties between ring-porous trees and diffuse-porous trees.

Quantitative PCR analysis of c-erb B-2 (HER2/neu) gene amplification and comparison with p185(HER2/neu) protein expression in breast cancer drill biopsies.

A PCR assay using capillary electrophoresis was designed for the detection of c-erbB-2 gene amplification in alcohol-formalin-acetic acid (AFA)-fixed, paraffin-embedded biopsies from 81 consecutive breast tumors. c-erbB-2 expression was analyzed in the same samples using immuno-histochemistry (IHC). In the competitive PCR assay, a single pTag plasmid containing a 4-nucleotide (nt)-deleted copy of a 124-nt sequence of c-erbB-2 and a 4-nt-deleted copy of a 120-nt sequence of GAPDH was co-amplified with genomic DNA extracted from 3 10-micrometer-thick tissue sections of the tumor biopsy. The percentage of tumor cells in the biopsy specimen and the percentage of tumor cells stained with the membrane anti-c-erbB-2 monoclonal antibody CB11 were recorded by a single pathologist on 2 consecutive sections. Among 81 consecutive tumor biopsies assayed by PCR, 21 (26%) displayed unequivocal c-erbB-2 amplification (actual gene copy number, AGCN > 4), 47 (58%) displayed no c-erbB-2 amplification (AGCN

In vivo induction of apoptosis in human lymphocytes by therapeutic fractionated total body irradiation.

Ionizing radiations have been reported as an in vitro apoptosis initiating stimulus in human lymphocytes. As the cytotoxicity of ionizing radiations and chemotherapeutic agents appears to be dependent on the efficacy of cell death induction, the manipulation of apoptosis initiation might be used as a means to supress some pathological process. In the present study the in vivo induction of gamma-ray mediated programmed cell death in humans is reported. The in vivo induction of apoptosis in peripheral blood lymphocytes (PBL) by ionizing radiations was investigated in 33 patients after each of two sessions (2 Gy and 4 Gy) of fractionated total body irradiation (FTBI) as part of their conditioning regimen before bone marrow transplantation. PBL committed to apoptosis were scored before irradiation (S1), 4 h (S2) and 24 h after 2 Gy (S3, 14-17 h after the second 2 Gy fraction). Nuclear morphology and chromatin-DNA were analysed by fluorescence microscopy immediately after blood sample withdrawal (I) and after 24 h in cell culture medium (II). When scored immediately after withdrawal, no circulating PBL with the apoptotic nuclear morphology were observed in S1 and S2 blood samples whereas S3 disclosed 21.9 +/- 11.7% of circulating lymphocytes with an apoptotic nuclear morphology. After 24 h in culture, S1 samples (before irradiation) generally contained less than 20% of apoptotic lymphocytes. A higher percentage of apoptotic cells was noted in some cases in relation with recent chemotherapy and possibly with pathology. After 24 h in culture, S2 and S3 samples contained 51.7 +/- 17.9% and 60.4 +/- 16.4% of apoptotic lymphocytes, respectively. These results confirm that ionizing radiations induce apoptosis in vivo in human lymphocytes and that the commitment to apoptosis can be determined after low doses (2 Gy) of therapeutic whole body irradiation. The results suggest that susceptibility to apoptosis induction by ionizing radiations could be related to previous therapy by cytotoxic drugs and possibly to the type of haematological malignancy.

[Superoxide dismutase (Cu/Zn) in cutaneous application in the treatment of radiation-induced fibrosis]. / Superoxyde dismutase (Cu/Zn) en application cutanée dans le traitement des fibroses radio-induites.

The aim of the paper consists to select criterias of the therapeutic effect of superoxide-dismutase (PEG-SOD) administrated as an ointment twice a day for 3 months. An original scoring method including qualitative and quantitative data was set up in order to appreciate the importance of the fibrosis and its variations when under local administration of PEG-SOD. Clinical and paraclinical controls were made as T0, T1 (6 weeks), T2 (3 months), T3 (6 months). After 6 months, results enabled us to show PEG-SOD in its galenic form was efficient on radiofibrosis with a 41% score reduction compared to pretreatment score T0, thus an improvement of nearly half of the potential theoretical recovery. The therapeutic efficiency was greater on the most recent fibrosis and there was a chronological order to the different recovery stages. After 6 weeks of administration pains were reduced or stopped; then after 3 months of treatment fibrous texture broke up and softened. An effective reduction of the surface as well as lightened of the pigmentation would not usually start until the 4th month after the start of treatment. PEG-SOD is thus an enzyme the therapeutic interest of which offers interesting prospects. The score which was set up enable a fair evaluation of the intensity and the variations of the treated fibrosis. A prospective study is currently on going in order to research the biological conditions in which the enzyme reacts.

[Immunoradionuclide localization of human neuroblastoma xenografted in nude mice using anti-GD2 labelled with I125]. / Localisation immunoscintigraphique de neuroblastomes humains greffés sur souris nude à l'aide d'anticorps anti-GD2 radio-iodés (125I).

Neuroblastoma is the most frequent tumour of the childhood under the age of 5. The staging and the follow up are achieved by MIBG scintigraphy, considered as the method of reference, but sometimes difficult to interpret . The availability of monoclonal antibodies against the ganglioside GD2, expressed on the cell membrane of neuroblastoma and neuro-endocrine cancers offers novel tools that deserve to be carefully explored. We investigated four mouse monoclonal antibodies (3 IgG3: BW704, 7A4, 60C3, and the IgG1 variant of BW704: MAK704), on nude mice xenografted with a human neuroblastoma (REM). Sixty one nude mice were included. The three former MAbs provided tumour imaging, the best results being obtained with BW704, followed by 7A4 and 60C3. MAK704 was disappointing. A control antiphosphorylcholine antibody (P51-1) did not give any tumour image in the three tested mice. Scintigraphy ratios tumour/liver and tumour/muscle reached 20 and 100 with BW704, respectively, on the 10th day. Good imaging quality was already obtained from the 24th h. The tumour uptake, calculated from radioactivity countings of resected samples, reached 22 +/- 3% of injected dose per gramme. These results let us hope that these antibodies could also provide highly contrasted images in humans and could open the way for therapeutic applications.

Strong uptake of 111In-pentetreotide by an MIBG-negative, xenografted neuroblastoma.

We investigated the distribution of 111In-pentetreotide (Octreoscan, Mallinckrodt) in nude mice xenografted with a human neuroblastoma cell line (SKLAN, derived from the LAN1 line). These cells develop into solid tumours in nude mice and can be grafted repeatedly in grafts of 10(8) cells. Animals were sequentially explored by scintigraphy 2, 4, 24 and 48 hr after i.v. injection of 2.5-4 MBq of the tracer and killed at various times up to 48 hr. 111In-pentetreotide was rapidly and strongly taken up by all tumours, with a tumour/muscle (T/M) ratio on resected samples of 20.0 +/- 5.7 at 2 hr, 23.7 +/- 3.0 at 4 hr, 75.6 +/- 12.6 at 24 hr and 78.7 +/- 12.4 at 48 hr, for tumours ranging from 0.5 to 8 g. Scintigraphy results were quantitatively in agreement. Pre-injection of a 15-20 times larger quantity of unlabelled octreotide s.c. reduced the tumour uptake by a factor of 2. For comparison, nude mice xenografted with the same cell line were also studied with 123I-MIBG (4 MBq). At 24 hr, the T/M ratio was 0.62 +/- 0.18. Two other cell lines (glioblastoma ROM and small-cell lung carcinoma SC41) which were similarly tested with 111In-pentetreotide (2.5-4 MBq) gave T/M ratios at 24 hr of 4.8 +/- 2.8 and 38.4 +/- 21.8, respectively. Pentetreotide seems to have a high affinity for this MIBG-negative neuroblastoma cell line, which exhibited a clearly higher tumour uptake than the 2 other lines. This work provides new experimental arguments in favour of the particular interest of somatostatin analogues in neuroblastoma and confirms our first clinical results.