Biomedical subjects

K Meflah

Publications and source records attributed to K Meflah.

At least 37 records · Page 2Linked to original sources

PubMed2000

Phase I trial of interleukin-2 and high-dose arginine butyrate in metastatic colorectal cancer.

INTRODUCTION: Interleukin-2 (IL-2) and sodium butyrate allow rats to be cured of peritoneal carcinomatosis from colon cancer. We performed a phase I trial of IL-2 and high-dose arginine butyrate (ArgB) in patients with advanced metastatic colorectal cancer. PATIENTS AND METHODS: From April to July 1997, six patients were included in the trail; they had a median age of 52 years, four had a performance status of 0, two had a performance status of 1 with normal biological functions. All patients had received at least two prior lines of chemotherapy. A fixed dose of 18 MIU/m2 IL-2,was administered by subcutaneous injection and ArgB was delivered via continuous intravenous infusion on days 1-6 with escalating doses starting at 2 g kg(-1) day(-1). RESULTS: The planned dose escalation was not possible because of toxicities. A daily ArgB dose of 2 g/kg was delivered for nine cycles. Level 2 (4 g/kg) could not be delivered in three of the six patients because of liver toxicity. The dose-limiting toxicities were fatigue and liver function disturbances. The maximum tolerated dose for ArgB was 3 g kg(-1) day(-1), in combination with IL-2 at 12 MIU m2 day(-1). No clinical response was seen. Pharmacokinetic analysis showed large intra- and interindividual variations. CONCLUSION: This schedule with a high dose of ArgB proved to be highly toxic with liver insufficiency. We will be running another trial with lower doses of ArgB calculated from the schedule used in the experimental model, starting at a dose of 20 mg kg(-1) day(-1) for ArgB and 200000 UI kg(-1) day(-1) IL-2, every 8 h.

Adjuvants, Immunologic↗

PubMed1999

Production and characterisation of a monoclonal antibody specific for apoptotic bodies derived from several tumour cell lines.

We have recently shown that apoptotic bodies (apobodies) derived from rat colon carcinoma cell lines (PROb) after sodium butyrate (NaB) treatment were able to cure rats with induced peritoneal carcinomatosis ( [BOISTEAU] ). A specific immune response was assumed to be involved since the serum of cured rats contained antibodies specific for apobodies. In the present study, a mAb (clone 6E8) produced by immunisation of rats with apobodies strongly recognized apobodies but had little reactivity with parental tumour cell lines, as demonstrated by enzyme-linked immunosorbent assay (ELISA), immunostaining and flow cytometry. Immunoelectron microscopy showed that 6E8 mAb mainly stained the hyaloplasm or cytosol of apobodies. A protein was detected at 67 kDa by immunoprecipitation of apobodies with mAb, followed by immunoblotting, using serum of rats immunised with apobodies. The 6E8 mAb recognized apobodies derived from several rat or human colon cancer cell lines and a rat glioma cell line, regardless of the apoptosis stimulus used (NaB, staurosporine or UV). Our results clearly show that 6E8 mAb defines an epitope specifically generated during apoptosis, which suggests that the protein recognized may be involved in the molecular cascade of apoptotic cell death.

Animals↗

PubMed1999

Antigen-presenting cells that phagocytose apoptotic tumor-derived cells are potent tumor vaccines.

We have reported recently that treatments combining injections of apoptotic bodies from tumor cells and interleukin 2 led to tumor regression and induced specific protection. In the present study, we show that tumor-bearing rats were cured with an 80% success rate by injection of antigen-presenting cells (APCs) that had phagocytosed apoptotic bodies derived from poorly immunogenic tumor cells, whereas phagocytic cells exposed to nonapoptotic tumor cell extracts were essentially without effect. In addition, curative vaccination using APCs that had phagocytosed apoptotic bodies generated a tumor-specific cytotoxic T-cell response and long-term protection from parental tumor challenge. Thus, systems using the processing and presentation of antigenic molecules by professional APCs after phagocytosis of apoptotic bodies appear to offer new possibilities for anticancer treatment.

Animals↗

PubMed1999

The C-terminus of bax is not a membrane addressing/anchoring signal.

It has been suggested that BCL-2 family members associate with certain organelles through their hydrophobic C-terminus which in the case of bcl-2, appears to play a key role in the regulation of apoptosis. We have investigated the association of bax with microsomal, nuclear and mitochondrial membranes using a cell-free system and found, contrary to bcl-2, that bax binds poorly to these organelles. Deletion of the C-terminal of bax (baxDeltaC) or exchanging the C-terminal ends of bax and bcl-XL suggests that the bax C-terminus is not an addressing/anchoring signal. In agreement with this observation, HL-60 cells transfected with either bax or baxDeltaC show no difference in sensitivity to an apoptotic signal. In the cell-free system, at low pH, bax becomes associated with mitochondria after a change of conformation, a result consistant with its structural homology with certain bacterial toxins. In HL-60 cells, as observed in the cell-free system, bax acquired a protease resistant conformation upon its translocation from the cytosol to the mitochondria after the induction of apoptosis.

Amino Acid Sequence↗

PubMed1999

Immunomodulatory effects of tumor-associated fibroblasts in colorectal-tumor development.

In order to elucidate the role of myofibroblasts in tumor development, we compared fibroblastic reactions and their implications in the immune response in progressive and regressive rat colorectal-tumor models. Immunohistochemical analyses revealed that T lymphocytes and monocytes/macrophages were found outside progressive tumors that were surrounded by a large sheath of myofibroblasts. In vitro experiments using fibroblast- vs. myofibroblast-containing collagen gels showed that the mechanical properties of these tumor-activated myofibroblasts prevent penetration of T lymphocytes and macrophages within tumor nodules. These results indicate that tumor-activated myofibroblasts may prevent physical contact between cancer-cells and immune cells, an essential phenomenon for effective destruction of cancer cells. Successful immunotherapy against cancer should therefore include complementary treatments against these tumor-associated fibroblasts.

1,2-Dimethylhydrazine↗

PubMed1999

Increased tumorigenicity of rat colon carcinoma cells after alpha1,2-fucosyltransferase FTA anti-sense cDNA transfection.

Accumulation of histo-blood group antigens such as Lewis b, Lewis Y and H increases tumor cell motility and tumorigenesis. Alpha1,2-fucosylation is a key step in the synthesis of these antigens. Two alpha1,2-fucosyltransferases, expressed in colorectal carcinomas, have been characterized (FUT1 and FUT2 in humans, FTA and FTB in rats). To define the relative contribution of each of these enzymes in tumor cell behavior, we have used an anti-sense transfection approach in rat colon carcinoma PROb cells, which synthesize mRNA encoding for both enzymes. We have previously reported that anti-sense transfection of a cDNA fragment of the FTB enzyme decreased H antigenic cell-surface levels and concomitantly decreased tumorigenicity. H antigens, detected by antibodies specific for H type 1, 3 or 4, were detected only on a splice variant of CD44 containing the product of exon v6. We now report the anti-sense transfection of an FTA cDNA fragment into PROb cells, which resulted in decreased enzymatic activity on a type 2 precursor and decreased cell-surface H type 2 antigen exclusively. Compared to controls, FTA anti-sense-transfected cells were significantly more tumorigenic in syngeneic animals but not in immunodeficient SCID mice. The UEA-I lectin, specific for H type 2, revealed that these structures were present on the CD44v6 variant and on an uncharacterized 80-kDa glycoprotein. Our results indicate that FTA and FTB fucosylate distinct glycan chains in the same cell, leading to opposite effects, under control of the immune system.

Animals↗

PubMed1999

Electron microscopy study of intrahepatic ultrasmall superparamagnetic iron oxide kinetics in the rat. Relation with magnetic resonance imaging.

Ultrasmall superparamagnetic iron oxide (USPIO) contrast agents for use in magnetic resonance imaging (MRI) are currently undergoing clinical evaluation. However, the images observed and the kinetic profiles obtained differ from one agent to another. In this study, BD IX rats received an intravenous penis injection of the USPIO contrast agents AMI-HS and AMI-227. A cytologic study of the liver was performed, and the data obtained were compared with those of MRI. Images acquired in light microscopy, transmission electron microscopy, microanalysis and electron diffraction provided data on the cell categories involved in the processing of these contrast agents, the importance and modalities of each category relative to this processing, and the modalities of agent elimination. AMI-HS was rapidly removed from the bloodstream by Kupffer's cells and hepatocytes and then eliminated through bile ducts. AMI-227 remained much longer in the blood compartment since it was processed very slowly by endothelial and Kuppfer's cells in the near absence of hepatocytic participation and thus of elimination by the bile ducts. These results allowed us to base our interpretation of MRI sequences on cytologic observations.

Animals↗

PubMed1999

T cell status influences colon tumor occurrence in min mice fed short chain fructo-oligosaccharides as a diet supplement.

We have previously shown that addition of short chain fructo-oligosaccharides (indigestible carbohydrates) to food prevented colon tumors in C57BL/6-Apc(Min/+) mice, a model for human colon cancer. As gut-associated lymphoid tissue was concomitantly developed, we suggested that the immune response generated by this food may interfere with carcinogenesis due to involvement of mucosal cells in the regulation of tissue homeostasis. In the present experiment, we tested whether T cell status may influence colon tumor formation in Min mice fed a food supplement of short chain fructo-oligosaccharides. Min mice depleted of CD4(+) and CD8(+) lymphocytes developed twice as many tumors as immunocompetent mice (0.8 as compared with 0.4, the mean number in 7-week-old Min mice when food supplementation began; P = 0.02). It is concluded that food supplementation with a substrate (a known prebiotic) fermented in the colon may stimulate a mechanism of immunosurveillance that would otherwise remain inefficient.

Animals↗

PubMed1999

Role of antigen-presenting cells in long-term antitumor response based on tumor-derived apoptotic body vaccination.

Cellular therapy prospects for cancer are based on the development of T cell response, resulting in efficient tumor rejection and long-term protection. We have previously shown that treatment combining injection of interleukin-2 and tumor-derived apoptotic bodies, but not tumor cell extracts, permits to reject parental tumor in 40% of rats. We observed the implication of antigen-presenting cells (APCs) and tumor-derived apoptotic bodies in the rejection of established peritoneal carcinomatosis. We demonstrated that apoptotic bodies could be efficiently phagocytosed by monocytes, triggering them to an APC phenotype. When using these phagocytosing APCs, derived from peritoneal or blood monocytes, the remission rate reached 80% of rats. However, due to the lack of specific markers of rat monocyte-derived cells, the precise role of APCs, dendritic cells and/or macrophages responsible for this therapeutic improvement remained to be clarified. In order to elucidate this question, we developed an in vivo preventive cellular therapy based on tumor-derived apoptotic bodies, where macrophages were either depleted or activated. We report here that in a preventive antitumoral apoptotic body vaccination that allows survival for 40% of treated rats, the antitumor response was characterized by a specific long-term memory (cured rats rejected a second parental tumor cell challenge). Depletion of resident macrophages with silica or clodronate liposomes appeared to promote apoptotic body vaccination efficiency, increasing the treatment to 66% of success. In this case, FACS analysis showed that peritoneal cells present are essentially immature APCs and freshly recruited NK cells. In contrast, the onset of peritoneal inflammation by thioglycollate, inducing massive recruitment and activation of macrophages, reduced the overall survival, whatever the treatment was. Also, even though the surviving rate was better in silica-treated rats than control, no long-term protection was elicited. Our data suggest that massive inflammation, recruiting numerous activated macrophages, could inhibit tumor antigen presentation by 'professional' APCs having phagocytosed apoptotic bodies, and defavor a specific antitumoral T cell response. Although effective responses were developed against parental tumor cells with silica/apoptotic body treatment, they seemed only partial, limited to primary cytotoxic efficiency. In conclusion, even if macrophages did not appear necessary for a primary response to tumor cells, these cells seemed to be implicated in the establishment of memory and long-term antitumor response.

Animals↗

PubMed1998

Interleukin-2/sodium butyrate treatment cures rats bearing liver tumors after acquired 5-fluorouracil resistance.

We studied the effect of immunotherapy using recombinant interleukin-2 (rIL-2) in combination with a differentiating agent, sodium butyrate (NaBut), on experimental 5-fluorouracil (5-FU)-resistant liver metastasis from colorectal cancer in rats. For this purpose, we used direct liver injection of 5-FU resistant cells, PRObRI, in syngeneic BDIX rats to establish liver tumors. The growth of liver metastasis was followed before and after NaBut/rIL-2 treatment by magnetic resonance imaging (MRI). The presence of liver tumors was checked by MRI 7 days after tumor cell injection. Evaluable rats were then assigned randomly to a control and an experimental group. The different treatments were started on day 10 and administered intraperitoneally (i.p.). Combined NaBut/rIL-2 treatment followed by MRI on days 56 and 91 was shown both to significantly reduce the growth of liver tumors and to prevent extrahepatic spread. In addition, NaBut/rIL-2 treatment induced a complete regression in 50% of the rats which remained free of disease.

Animals↗

PubMed1998

Potentiation of apoptosis by mitochondria in a cell-free system.

Using a cell-free system, we show that rat liver mitochondria, but not mitochondrial extracts, potentiated apoptosis triggered by cytosols derived from apoptotic cells. Apoptosis potentiated by mitochondria appeared to be inhibited by caspase 3 but not by caspase 1 inhibitors. A cytosolic caspase-3-like activity was increased by the addition of mitochondria to apoptotic cytosols; the latter activation was inhibited by the addition of bcl-2. Chelation of calcium by EGTA significantly and specifically inhibited the apoptosis potentiated by mitochondria as well as the increase of caspase-3-like activity. The incubation of mitochondria with apoptotic cytosols led to the release of cytochrome c, this latter phenomenon being inhibited by EGTA. Calcium or cytochrome c and dATP, however, did not reproduce the mitochondrial potentiation in the absence of the organelle. Thus, mitochondria can initiate and potentiate apoptosis through similar but not identical mechanisms.

Animals↗

PubMed1998

Induction of a caspase-3-like activity by calcium in normal cytosolic extracts triggers nuclear apoptosis in a cell-free system.

Calcium is involved in several steps of the apoptotic process. In nuclei, endonucleases are presumed to be the main targets of calcium; however, little is known about its role during the cytosolic phase of apoptosis. We used a cell-free system to address this question. Our results show that CaCl2 triggered nuclear apoptosis (i.e. typical morphological change and DNA fragmentation) at concentrations of 5 mM. This concentration was lowered 10-fold by the co-incubation with cytosolic extracts from nonapoptotic cells. Apoptotic changes induced by the incubation of nuclei with CaCl2 in the presence of these cytosols were strongly reduced in the presence of an inhibitor of caspase-3 and to a lesser extent by an inhibitor of caspase-1. We also show that calcium-induced apoptosis is affected by protease inhibitors such as N-tosyl-L-phenylalanine chloromethyl ketone, but not by calpain or several lysosomal protease inhibitors. The addition of CaCl2 to the cell-free system increased a caspase-3 activity in nonapoptotic cytosols as shown by specific antibodies and an enzymatic assay. No activation of a caspase-3-like activity by the addition of cytochrome c was observed in these extracts under similar conditions. The enhanced caspase-3 activity induced by calcium was inhibited by protease inhibitors affecting morphological nuclear apoptosis except for those responsible for the degradation of lamin A. These results suggest that CaCl2 could trigger, in normal cells, an apoptotic cascade through the activation of cytosolic caspase-3 activity.

Amino Acid Sequence↗

PubMed1998

Effect of resistant starch and/or fat-soluble vitamins A and E on the initiation stage of aberrant crypts in rat colon.

This study reports the modulating effects of resistant starch (RS) and the fat-soluble vitamins A or E, alone or in combination, on initiation of preneoplastic lesions in rat colon aberrant crypt foci (ACF) induced by 1,2-dimethylhy-drazine. One group of male Sprague-Dawley rats was fed a basic diet and five groups were fed experimental diets supplemented with 25% RS, 200 IU vitamin A, 5 IU vitamin E, 25% RS + 200 IU vitamin A, or 25% RS + 5 IU vitamin E for four weeks. After induction by 1,2-dimethylhydrazine, all the animals were fed basic diets for four more weeks before sacrifice. Compared with the basic diet, only the vitamin A-supplemented diet significantly reduced the incidence of ACF. The vitamins incorporated in the animals' diets increased the vitamin concentrations in hepatic and colonic cells compared with the animals fed the basic diet. The preventive effect of vitamin A seems to be due to a direct effect on colonic epithelial cells. The three diets supplemented with RS significantly decreased cecal pH and bacterial beta-glucuronidase activity and increased cecal weight and fecal output. The retrograde high-amylose maize, type 3, used in this study does not significantly decrease ACF. This RS has an effect on the colon similar to that of nonstarch polysaccharides. Neither biochemistry nor four weeks of dietary supplementation is likely sufficient for adaptation of the rat colonic flora.

1,2-Dimethylhydrazine↗

PubMed1998

Synergistic effects of relaxation and susceptibility in differentiation between compartmentalized and noncompartmentalized tissues.

RATIONALE AND OBJECTIVES: This study illustrates the synergistic effects of relaxation- and susceptibility-based contrast enhancement. Using a combination of gadolinium (Gd) and dysprosium (Dy) complexes and a sequence capable of taking advantage of the particular relaxation behavior of this combination, the difference between compartmentalized and noncompartmentalized regions was significantly enhanced. METHODS: Magnetic resonance imaging of the rat kidney was performed before and immediately after the administration of a combination of Gd and Dy chelates (Gd-DTPA-BMA and Dy-DTPA-BMA). RESULTS: The signal intensity (SI) of the renal parenchyma was reduced by 85%, whereas the collecting tubes had a 100% increase of their SI as demonstrated by a short repetition time (600 msec), long echo time (50 msec), and spin-echo sequence. CONCLUSIONS: The high R2* effect, specific to the compartmentalized tissues, associated with the moderately high R1 and R2 developed in the remaining areas, results in an important improvement in tissue differentiation, which potentially is useful for the evaluation of pathological changes as in tubular necrosis.

Animals↗

PubMed1997

5-Fluorouracil-resistant colonic tumors are highly responsive to sodium butyrate/interleukin-2 bitherapy in rats.

Advanced colorectal cancer is generally refractory to 5-fluorouracil (5-FU) chemotherapy. This is linked to the emergence of resistant cell populations, probably due to a selection process. The identification of molecular markers and the improvement of alternative therapies thus remain important. We have used as an experimental model a rat colon cancer cell line (PROb), which exhibits features similar to those of the human situation. 5-FU treatment of rats bearing PROb tumors enhanced their survival but did not lead to cure. A PROb 5-FU-resistant subline (PRObR1) was obtained by continuous in vitro exposure to 5-FU. Resistance to 5-FU was accompanied by a 2-fold increase in thymidylate synthase activity and a substantially higher incorporation of thymidine in the presence of 5-FU, compared with parental PROb cells. Unexpectedly, in syngeneic rats, PRObR1 tumors exhibited delayed growth when compared with parental PROb tumors. This was ascribed to an increased sensitivity of PRObR1 cells to host immune response since no growth delay was observed in immunocompromised nude mice and since there was no detectable difference in proliferation rates between PROb and PRObR1 cells. 5-FU treatment was inefficient in prolonging the survival of rats bearing PRObR1 tumors. In contrast, an immunotherapeutic protocol combining sodium butyrate and recombinant interleukin-2 (NaBut/rIL-2) cured 80% of the rats bearing established PRObR1 tumors. Our results suggest that NaBut/rIL-2 treatment is efficient against 5-FU-chemoresistant rat colon cancer.

Animals↗

PubMed1997

Short-chain fructo-oligosaccharides reduce the occurrence of colon tumors and develop gut-associated lymphoid tissue in Min mice.

C57BL/6J-Min/+ mice, which are heterozygous for a non-sense mutation in the Apc gene, provide a model for both familial adenomatous polyposis and sporadic colon cancers. In our study, gut tumors and small intestine lymphoid nodules were counted in Min mice fed fiber-enriched diets for 6 weeks. Neither starch-free wheat bran nor resistant starch modified the number of tumors. However, short-chain fructo-oligosaccharides dramatically reduced the incidence of colon tumors and concomitantly developed gut-associated lymphoid tissue. Our experiment shows that short-chain fructo-oligosaccharides counteract advanced stages of colon carcinogenesis, possibly via stimulation of antitumoral immunity by modulation of the colonic ecosystem.

Animals↗

PubMed1997

Apoptosis induced by sodium butyrate treatment increases immunogenicity of a rat colon tumor cell line.

We have recently demonstrated that a treatment combining the cell differentiating agent sodium butyrate (NaBut) and interleukin-2 (IL2) resulted in a remission of established peritoneal colorectal carcinomatosis in rats. NaBut or IL2 treatment alone, never cured these tumour-bearing rats. In the present investigation, we report that NaBut-treatments induce apoptosis in the colonic cancer cells both in vitro and in vivo. We postulated that the significant therapeutic effect of NaBut/IL2 treatment can be mainly attributed to a NaBut-induced apoptosis of the tumoural cells increasing their immunogenicity. Indeed, treatment which combined apoptotic bodies (apobodies) as cell vaccine, plus IL2 immunotherapy significantly increased tumour remission and survival rate of the vaccinated rats, whereas IL2 treatment alone did not. We observed that the cured rats presented long-term protection against subsequent challenge with the parental tumour cells. This latter result suggests that these treatments generate an immune protection. This was confirmed by the presence, in the sera of the cured rats, of anti-tumoural antibodies directed against both the apobodies and the tumour cells, but not against normal colonocytes. In addition, we show that injections of apobodies before administration of the parental tumour cells results in a partial protection. We provide the first evidence that apobodies, derived from cancer cells after NaBut-treatment, induce a specific immune response against parental tumours cells. These data suggest that the distinctive immunologic properties of apobodies could provide a valuable tool in colorectal cancer immunotherapy.

Journal Article↗

PubMed1997

Effects of resistant starch- and vitamin A-supplemented diets on the promotion of precursor lesions of colon cancer in rats.

We have evaluated the potential protective effect of resistant starch (RS)- and vitamin A-supplemented diets on the promotion of preneoplasic lesions of rat colon, aberrant crypt foci (ACF), induced by 1,2-dimethylhydrazine dihydrochloride (DMH). We have tried to show whether the association of these two dietary constituents in the same diet could have synergistic effects. RS, vitamin A, and RS+ vitamin A were incorporated into the rat diets. Experimental diets were given one week after DMH injection and maintained for 12 weeks until the animals were sacrificed. The total number of ACF decreased with the three experimental diets. For RS- and RS + vitamin A-supplemented diets, this decrease is primarily due to a decrease in small ACF. For the vitamin A-supplemented diet, small and large ACF have a tendency to decrease. The effects of the diets on parameters influencing colon carcinogenesis were also studied. Only RS- and RS + vitamin A-supplemented diets have modified cecal pH, fecal and cecal butyrate contents, fecal excretion, cecal weight, and colon length. Vitamin A has been observed in colonic epithelial cells of rats receiving vitamin A- and RS+ vitamin A-supplemented diets. The association between RS and vitamin A shows neither a cumulative nor a synergistic protective effect.

Animals↗