Effects of a new triazinoaminopiperidine derivative on adriamycin accumulation and retention in cells displaying P-glycoprotein-mediated multidrug resistance.

A new triazinoaminopiperidine derivative, Servier 9788 (S9788), was investigated for its ability to increase Adriamycin (ADR) accumulation and retention in two rodent (P388/ADR and DC-3F/AD) and three human (KB-A1, K562/R and COLO 320DM) cell lines displaying the P-glycoprotein (P-gp)-mediated multidrug resistance (MDR) phenotype. Depending on the cell line S9788 was shown to be two to five times more active and five to 15 times more potent than Verapamil (VRP) in increasing ADR accumulation in resistant cells. ADR retention in KB-A1 cells maintained in a concentration of 10 microM S9788 was twice that in VRP-treated cells, and similar to that measured in the untreated sensitive KB-3-1 cells. Although 5 microM S9788 and 50 microM VRP gave the same values of ADR uptake in KB-A1 cells, S9788 was shown to induce a greater ADR retention following cell wash and post-incubation in resistance modifier- and ADR-free medium. Taking into account that S9788 had no effects on ADR accumulation and retention in sensitive KB-3-1 cells, it can be suggested that S9788 inhibits specifically the P-gp dependent ADR efflux, and in a manner less reversible than that observed with VRP. Moreover, [3H]azidopine photolabeling of P-gp, in P388/ADR plasma membranes, was completely inhibited by 100 microM S9788. Although S9788, as VRP, had no effect on the cell cycle of P388 cells, 5 microM S9788 increased 700-fold the efficacy of ADR to block P388/ADR cells in the G2+M phase of the cell cycle. Together, these results show that the sensitization, by S9788, of cell lines resistant to ADR is mainly due to an increase in ADR accumulation and retention, leading to an increase in the number of resistant cells blocked in the G2+M phase.

Integrated system for the screening of the specificity of protein kinase inhibitors.

Tyrosine protein kinases (TPKs) play a major role in the transformation of cells. They are currently used as molecular targets for new generations of anticancer compounds. Numerous TPKs have been described from various tissues using either classical molecular biochemical techniques or cloning strategies. As a natural extension of these discoveries, a large number of "specific" inhibitors have been described in the literature. The major problem with these inhibitors is that there is no simple way to compare their specificity and/or selectivity from one report to another. We have set up a simple, straightforward technique to compare the inhibitory potency of 14 classical inhibitors towards six well-described and at least partially purified protein kinases. This technique is based on a new assay, easy to carry out and non-restrictive in terms of the type of protein substrate used. It permits direct comparisons between the results obtained from various sources. Data obtained showed that, when assessed in this integrated system, specificity and selectivity of many "classical" inhibitors are often weak, thus demonstrating that a universal technique such as ours is essential for the molecular screening of new protein kinase inhibitors. Compounds showing specificity for this panel of protein kinases will be more easily targeted to some defined types of oncogene and of transformed cells.

Multidrug-resistance circumvention and inhibition of [h-3] azidopine photolabeling of p-glycoprotein by new dihydropyridine derivatives displaying a low-affinity for calcium channels.

This study was aimed to characterize the reversing activity of S16209 and S16317, two new dihydropyridines with low affinity for calcium channels. In vivo, S16209 (75 mg/kg) and S16317 (25 mg/kg) potentiate the antitumor activity of vincristine (VCR) in VCR-resistant leukemia bearing mice. In vitro, a complete sensitization to adriamycin (ADR) or VCR is obtained with 2.5 muM of S16209 in S1/tMDR and KB-A1 cells and with 2.5 muM of S16317 in S1/tMDR and P388/ADR-10 cells. These two compounds are also more potent than verapamil and cyclosporin A in increasing actinomycin-D cytotoxicity in DC-3F/AD cells. In the presence of ADR or VCR, a 4 h co-incubation followed by a post-incubation of 20 h with 2.5 muM S16209 is sufficient to completely overcome the resistance of human KB-A1 and S1/tMDR cells to these cytotoxic drugs. S16209 and S16317 increase ADR accumulation in resistant cells, and completely inhibit the photolabeling of P-gp by [H-3]azidopine at 100 and 10 muM, respectively, suggesting that the reversing activity of these two compounds is mainly due to a specific inhibition of the P-gp mediated efflux of cytotoxic drugs.

A zinc chelator inhibiting gelatinases exerts potent in vitro anti-invasive effects.

Matrix metalloproteinases are zinc metalloenzymes involved in remodelling of the extracellular matrix. We compared the anti-invasive properties of a zinc ejector matrix metalloproteinase inhibitor with those of reference compounds (hydroxamic acid-based BB-94 and Ro-31-9790) which form inactive ternary complexes with the enzymes and the catalytic zinc. We show that the compound undecadenedioic acid bis-[[2-(3 H-imidazol-4-yl)-ethyl]-amide] (S 30372) is active against gelatinases, chelates zinc and exhibits enzymatic features compatible with the potential to extract zinc from gelatinases. We then used five invasive cell lines in the Matrigel invasion chamber assay (NIH-3T3 fibroblasts, Lewis lung carcinoma cells, EJ138 and J82 bladder carcinoma and HT1080 fibrosarcoma cells). With the exception of J82 cells which were unaffected by the three inhibitors, all remaining cells were substantially more sensitive to S 30372 in terms of maximal inhibition of invasion attained. This suggests that matrix metalloproteinase inhibitors with zinc chelating/ejecting properties may be more efficient in preventing tumor progression.

Limb salvage by neoadjuvant isolated perfusion with TNFalpha and melphalan for non-resectable soft tissue sarcoma of the extremities.

AIMS: Patients with non-resectable soft tissue sarcomas of the extremities do not live longer if they are treated by amputation or disarticulation. In order to avoid major amputations, we tested isolated limb perfusion (ILP) with tumour necrosis factor alpha (TNF)+melphalan+/-interferon-gamma (IFN) as a pre-operative, neoadjuvant limb salvage treatment. METHODS: Twenty-two patients were included (six men and 16 women; three upper limb and 19 lower limb tumours). The AJCC stage was IIA in four patients, III in seven and IV in 11. Thirteen cases were recurrent or progressive after previous therapy; five tumours had a diameter >/=20 cm, and four were multiple or regionally metastatic. There were six malignant fibrous histiocytomas, five liposarcomas, four malignant peripheral nerve sheath tumours, three rhabdomyosarcomas, two leiomyosarcomas, one recurrent extraskeletal osteosarcoma and one angiosarcoma. RESULTS: Twenty-four ILPs were performed in the 22 patients, and 18 (82%) experienced an objective response: this was complete in four (18%) and partial in 14 (64%). Three patients had a minimal or no response and the tumour progressed in one case. All patients had fever for 24 hours but only one developed a reversible grade 3 distributive shock syndrome with no sequelae. There was no grade 4 toxicity. Seventeen patients (77%) underwent limb-sparing resection of the tumour remnants after a median time of 3.4 months: 10 resections were intracompartmental and seven extracompartmental. Surgery included flaps or skin grafts in five patients, arterial replacement in two and knee arthrodesis in one. Adjuvant chemotherapy was given to eight patients and radiotherapy to six. In one patient amputation was necessary after a second ILP. Secondary amputations were performed for recurrence in two patients, resulting in an overall limb salvage rate of 19/22 (86%). After a median follow-up of 18.7 months, 10 recurrences were recorded: seven were both local and systemic and three were only local. The median disease free and overall survival times have been >12.5 and 18.7 months respectively: this is similar to the outcome after primary amputations for similar cases. CONCLUSION: ILP with TNF and chemotherapy is an efficient limb sparing neoadjuvant therapy for a priori non-resectable limb soft tissue sarcomas.

Use of the main tyrosine protein kinase activity purified from HL-60 in the search for a new class of anticancer compounds.

A tyrosine protein kinase (TPK) partially purified and two co-purified proteins (pp39 and pp44) from HL-60 cells were used as a basis for a SDS-PAGE electrophoresis screening test of inhibitors of the activity catalyzed by this enzyme. Such inhibitors may constitute a new class of anticancer agents. TPK is a protein of about 32 to 35 kDa as measured by gel-sizing exclusion. This enzyme is apparently the main tyrosine protein kinase from the cytosol of HL-60 cells. The test was assessed using known inhibitors of various types of protein kinases. Erbstatin and Cibcron Blue--a nucleotide analog--were shown to be potent inhibitors of TPK activity.

Studies of the potency of protein kinase inhibitors on ATPase activities.

Tyrosine as well as serine/threonine protein kinase inhibitors have potentially two sites of interaction with their targets: the protein-substrate binding site and the ATP binding site. The latter could be modelized by measuring the capacity of protein kinase inhibitors to inhibit ATPase activities. In order to do so, we assess a novel, highly sensitive HPLC method based on hydrophilic separation of [gamma-32P]ATP and [32P]Pi. The novel assay is presented. Furthermore, the potency of 13 protein kinase inhibitors was tested on two types of ATPase, namely: apyrase and partially purified liver mitochondria F1-ATPase. The method described for the assay of ATPase can be used with almost any type of enzyme catalyzing this activity. Only cibacron blue and suramin show interesting capacities in inhibiting these ATPase activities pointing out that several widely used protein kinase inhibitors are at least somewhat specific in that they do not inhibit these two ATPases.

Resection of metastases to the alimentary tract from malignant melanoma.

Metastases to the celio-mesenteric organs from malignant melanoma are studied retrospectively in 22 autopsied cases and in seven surgically treated patients. Metastatic disease to one or more celio-mesenteric organs was found in 86.3% of postmortem examinations: the liver was the most frequently involved organ, followed by the pancreas, peritoneum, small bowel, biliary tract spleen, colon and stomach, in that order. Surgery only resulted in long-term asymptomatic survival in those patients with a long relapse-free interval and a single metastasis. It is concluded that, apart from cases with intestinal obstruction or massive GI bleeding, resection should be reserved for patients with a relapse-free interval of at least 15 months and with a single "alimentary" metastasis. It is suggested that diagnosis of systemic spread at the asymptomatic stage would improve prognosis of stage IV malignant melanoma.

[Evaluation of a school health program for high school and gymnasium students in the canton of Vaud: methodological tools]. / L'évaluation d'un programme de santé scolaire pour les apprentis et les gymnasiens vaudois: outils méthodologiques.

A pilot health school programme has been set up on an experimental way since January 1984 in a few professional and high schools of the Canton of Vaud. The paper presents the methodological tools that will allow an evaluation of various aspects of this experience, which has lasted for three years. The process will be evaluated among three populations: the pupils, the teachers and the members of the health team. The results will be evaluated in two fields: the screening of hypertension and tuberculosis on one side and the outcome of individual counseling on the other side. The paper illustrates both the utility and the difficulties linked to the setting up of such an evaluation process: it underlines the complexity of choosing indicators that are useful accessible and acceptable among the huge number of available data.

[Splenic abscess: therapeutic options]. / Abcès spléniques: options thérapeutiques.

Pyogenic splenic abscess is a rare condition that tends to occur in patients with underlying often infectious diseases. The sensitivity of ultrasonography and computerized tomography has improve early diagnosis and follow-up. In our hands, splenectomy remains the therapy of choice, but we report the resolution of two uncomplicated splenic abscesses with antibiotics alone.

[Withdrawal of a drug from the market for safety reasons: process and communication]. / Le retrait d'un médicament pour raison de sécurité: Processus et communication.

Each year, drugs must be withdrawn from the market following the identification of serious adverse reactions. We report here on an enquiry addressed to persons involved in the recent withdrawal of nefazodone, and a survey of physicians and pharmacists. Our results show the complexities of such withdrawal decisions and the difficulty in communicating them through the network of health professionals down to the patients.

[Liver transplantation with partial clamping of the inferior vena cava: preliminary experiences]. / Transplantation hépatique avec clampage partiel de la veine cave inférieure: premières expériences.

Caval reconstruction during orthotopic liver transplantation is usually achieved by two end-to-end anastomoses and extracorporeal veno-venous bypass. We report our preliminary experience with a new technique by lateral clamping of the recipient's cava and a side-to-side caval anastomosis without by-pass. This new technique appears safe and shortens the operative time.

[Acute arterial obstructions of the limbs in the aged]. / Obstructions artérielles aiguës des membres chez les vieillards.

131 cases of acute arterial obstruction were treated during 1973-1977, 70 (52%) of these cases were seen in patients of over 70 years of age and 20 out of them died during the first two months of the surgical treatment, 8 patients were amputed. The long term evaluation of 40 patients who survived for more than two months after the surgical treatment shows a very high mortality (14 of the 40 surviving patients died during the first two years of the surgical treatment). Satisfactory results were obtained in 18 patients. The long term results after arterial obstruction in old patients has high mortality and the number of amputations is directly proportional to the delay before commencement of the surgical treatment.

[External fixator in severe traumatic injuries of the pelvis: results apropos of 20 consecutive cases]. / Le fixateur externe dans les lésions traumatiques graves du bassin: résultats à propos de 20 cas consécutifs.

From August 91 to December 94, 20 external fixators were used for severely injured patients (avg. ISS 25.2). The fractures were essentially open book with or without lateral compression and vertical lesions. The indication for fixation was treatment of shock and stabilization in 8 cases, stabilization alone in 9 cases, and in 3 cases as complementary fixation after internal fixation of posterior lesions. The fixation of the pelvis was effective on the amount of blood loss. One acetabulum fracture required surgery, two patients had internal fixation for loss of reduction and two others for late pubic and posterior pain. The clinical results are good; they are more related to the severity of the initial lesion than to the mode of fixation or the quality of the reduction. No superficial sepsis or osteitis was observed in relation to the pins.

[Cure of recurrent ascites in alcoholic hepatitis following transient placement of a LeVeen shunt]. / Guérison de l'ascite réfractaire dans l'hépatite alcoolique, après mise en place transitoire d'un shunt de LeVeen.

Three cases of severe alcoholic hepatitis associated with refractory ascites are reported. A LeVeen shunt was inserted which was effective and well tolerated. However, the shunt was removed in all 3 patients 8 months, 12 months and 9 years after insertion because of spontaneous migration (n = 2) or superior vena cava thrombosis (n = 1). At time of shunt removal all 3 patients had micronodular cirrhosis, but none had recurrent ascites during the 1-3 years of observation and despite the absence of diuretics. The eventual transient aspect of refractory ascites associated with alcoholic hepatitis is discussed, together with the potential beneficial and temporary role of LeVeen shunts in this condition.