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ABSTRACT: 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 </= 2) and 7 (9%) could not be analyzed due to an insufficient amount of DNA. Six samples (7%) were considered inconclusive since the percentage of tumor cells was <20%. Analysis of c-erbB-2 expression by IHC showed that among the 21 amplified specimens 15 displayed strong staining, while all non-amplified samples (47) displayed no or only weak staining. The concordance of the 2 techniques was 91%. We conclude that c-erbB-2 gene amplification can be accurately quantitated by competitive PCR performed on small, fixed and embedded tumor samples.
International Journal of Cancer 11/1999; 83(2):157-61. · 5.44 Impact Factor
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ABSTRACT: 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.
British Journal of Radiology 10/1995; 68(813):997-1003. · 1.31 Impact Factor
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ABSTRACT: 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.
Bulletin du cancer 09/1994; 81(8):659-69. · 0.67 Impact Factor
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ABSTRACT: 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.
Bulletin du cancer 08/1994; 81(7):593-8. · 0.67 Impact Factor
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ABSTRACT: 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.
International Journal of Cancer 05/1994; 57(2):245-6. · 5.44 Impact Factor
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http://dx.doi.org/10.1051/radiopro/1992022.
