YAP/TEAD involvement in resistance to paclitaxel chemotherapy in lung cancer.

The Yes-associated protein (YAP) oncoprotein has been linked to both metastases and resistance to targeted therapy of lung cancer cells. We aimed to investigate the effect of YAP pharmacological inhibition, using YAP/TEA domain (TEAD) transcription factor interaction inhibitors in chemo-resistant lung cancer cells. YAP subcellular localization, as a readout for YAP activation, cell migration, and TEAD transcription factor functional transcriptional activity were investigated in cancer cell lines with up-regulated YAP, with and without YAP/TEAD interaction inhibitors. Parental (A549) and paclitaxel-resistant (A549R) cell transcriptomes were analyzed. The half-maximal inhibitory concentration (IC50) of paclitaxel or trametinib, which are Mitogen-Activated protein kinase and Erk Kinase (MEK) inhibitors, combined with a YAP/TEAD inhibitor (IV#6), was determined. A three-dimensional (3D) microfluidic culture device enabled us to study the effect of IV#6/paclitaxel combination on cancer cells isolated from fresh resected lung cancer samples. YAP activity was significantly higher in paclitaxel-resistant cell lines. The YAP/TEAD inhibitor induced a decreased YAP activity in A549, PC9, and H2052 cells, with reduced YAP nuclear staining. Wound healing assays upon YAP inhibition revealed impaired cell motility of lung cancer A549 and mesothelioma H2052 cells. Combining YAP pharmacological inhibition with trametinib in K-Ras mutated A549 cells recapitulated synthetic lethality, thereby sensitizing these cells to MEK inhibition. The YAP/TEAD inhibitor lowered the IC50 of paclitaxel in A549R cells. Differential transcriptomic analysis of parental and A549R cells revealed an increased YAP/TEAD transcriptomic signature in resistant cells, downregulated upon YAP inhibition. The YAP/TEAD inhibitor restored paclitaxel sensitivity of A549R cells cultured in a 3D microfluidic system, with lung cancer cells from a fresh tumor efficiently killed by YAP/TEAD inhibitor/paclitaxel doublet. Evidence of the YAP/TEAD transcriptional program's role in chemotherapy resistance paves the way for YAP therapeutic targeting.

CXCR4 inhibitors could benefit to HER2 but not to triple-negative breast cancer patients.

The CXCR4 receptor and its ligand CXCL12 (also named stromal cell-derived factor 1, SDF1) have a critical role in chemotaxis and homing, key steps in cancer metastasis. Although myofibroblasts expressing CXCL12 are associated with the presence of axillary metastases in HER2 breast cancers (BC), the therapeutic interest of targeting CXCR4/CXCL12 axis in the different BC subtypes remains unclear. Here, we investigate this question by testing antitumor activity of CXCR4 inhibitors in patient-derived xenografts (PDX), which faithfully reproduce human tumor properties. We observed that two CXCR4 inhibitors, AMD3100 and TN14003, efficiently impair tumor growth and metastasis dissemination in both Herceptin-sensitive and Herceptin-resistant HER2 BC. Conversely, blocking CXCR4/CXCL12 pathway in triple-negative (TN) BC does not reduce tumor growth, and can even increase metastatic spread. Moreover, although CXCR4 inhibitors significantly reduce myofibroblast content in all BC subtypes, they decrease angiogenesis only in HER2 BC. Thus, our findings suggest that targeting CXCR4 could provide some therapeutic interest for HER2 BC patients, whereas it has no impact or could even be detrimental for TN BC patients.

[Anticoagulant therapy with recombinant hirudin in patients with thrombopenia induced by heparin]. / Traitement anti-thrombique par hirudine recombinante chez des patients ayant une thrombopénie induite par l'héparine.

OBJECTIVES: Heparin-induced thrombocytopenia is an uncommon and severe complication of heparin therapy. Both venous and arterial thromboembolic events can occur, requiring withdrawal of the heparin therapy. When anticoagulant therapy is mandatory, recombinant hirudin can be used. METHODS: We used recombinant hirudin (HBW 023) in 6 patients with heparin induced thrombocytopenia. In case of venous thromboembolism, an initial intravenous bolus (0.07 mg/kg) was followed by continuous infusion (0.05 mg/kg/h); for arterial thromboembolism the initial bolus was 0.7 mg/kg and infusion rate 0.15 mg/kg/h. When possible oral anticoagulants were started and hirudin withdrawn when the INR ratio reached 3. RESULTS: The clinical course was uneventful in all 6 patients. There was no recurrent thromboembolism. Cephalin-activated coagulation time (patient/control) varied between 1.8 and 3.5 (median 2.4) during hirudin administration. Platelet count rose to the nadir (median 70 x 10(9)/l, range 15-90) reaching over 100 x 10(9)/l in all patients between the third and sixth day (median 5 days) after stopping heparin. CONCLUSION: Intravenous administration of hirudin provides effective immediate anticoagulation in patients with heparin-induced thrombocytopenia, thus allowing conversion to oral anticoagulants without risking recurrent thromboembolism.

Use of recombinant hirudin as antithrombotic treatment in patients with heparin-induced thrombocytopenia.

Heparin-induced thrombocytopenia is a rare but severe complication of heparin therapy that can result in severe venous or arterial thromboembolic events and whose treatment remains partially unanswered. Recombinant hirudin is potentially effective as an antithrombotic treatment in the management of heparin-induced thrombocytopenia, given its potent antithrombin effects without known interaction with platelets. We report the results obtained with intravenous recombinant hirudin (HBW 023) administered on a compassionate basis to patients suffering from heparin-induced thrombocytopenia. Six patients suffering from heparin-induced thrombocytopenia were submitted to intravenous recombinant hirudin (HBW 023) administered at a dose of 0.05 mg/kg/hr after an initial bolus injection of 0.07 mg/kg in the case of a venous thromboembolic event, and at a dose of 0.15 mg/kg/hr with the same initial bolus injection in the case of an arterial thromboembolic event. Whenever possible, oral anticoagulation with acenocoumarol was introduced at the same time as recombinant hirudin, which was interrupted as soon as the international normalized ratio reached 3. Clinical events, particularly thromboembolism and bleeding, were noted; activated partial thromboplastin time (aPTT), and platelet count were assessed throughout the administration of recombinant hirudin. Heparins responsible for heparin-induced thrombocytopenia were porcine sodium or calcium heparinate in four cases, nadroparin in one case, and enoxaparin in one case. Thrombocytopenia was discovered on routine systematic platelet count in two patients and after the occurrence of arterial and venous thromboembolism in two patients, respectively. After discontinuation of heparin and the onset of recombinant hirudin, clinical evolution was uneventful in all patients, with no recurrence of thromboembolism, limb amputation, or hemorrhagic complication. The aPTT ratio varied from 1.8 to 3.5 (median 2.4) throughout administration of recombinant hirudin. Platelet count rose from nadir (median value 60 x 10(9), 15 to 90) to above 100 x 10(9)/L in every patient within 3-6 days (median 5), after discontinuation of heparin. Intravenous administration of recombinant hirudin ensured safe anticoagulation in patients with heparin-induced thrombocytopenia and made it possible to wait for oral anticoagulation to become efficient and platelet count to return to normal values without occurrence or recurrence of thromboembolism.

[Subcutaneous recombinant hirudin in the treatment of deep venous thrombosis. A pharmacokinetic study]. / Hirudine recombinée par voie sous-cutanée dans le traitement de la thrombose veineuse profonde constituée. Etude pharmacocinétique.

Recombinant hirudin (HBW 023) has a pure and specific antithrombotic activity. It could be more effective than heparin in the treatment of deep venous thrombosis. Its half life is about three hours when administered intravenously which requires continuous infusion whereas subcutaneous administration can ensure stable plasma concentrations and antithrombotic activity over a period of approximatively 12 hours. The aim of the study was to check the safety and clinical and radiographic efficacy of recombinant hirudin administered subcutaneously to patients with recent deep venous thrombosis and to analyse the pharmacokinetics of the product and its effects on tests of coagulation. Ten patients were treated with 0.75 mg/kg of subcutaneous recombinant hirudin twice a day for 5 days. Anticoagulation was performed with standard heparin and acenocoumarol. Bilateral phlebography, pulmonary angiography or ventilation and perfusion scintigraphy were carried out before and on the 5th day of recombinant hirudin treatment. The activated cephalin time and standard anticoagulant tests and the plasma kinetics of recombinant hirudin were assayed between the 1st and 12th hour on the first and fifth days of treatment. The clinical course was simple in all but one patient who had a recurrence of pulmonary embolism on the 4th day justifying thrombolytic treatment. No haemorrhagic complications or secondary biological effects were observed. On the 5th day, control phlebography was unchanged or improved in all patients. The peak plasma concentration of recombinant hirudin was observed between the 3rd and the 4th hour following subcutaneous injection. The activated cephalin time was increased in parallel with increased concentrations of recombinant hirudin.(ABSTRACT TRUNCATED AT 250 WORDS)

A pilot study of subcutaneous recombinant hirudin (HBW 023) in the treatment of deep vein thrombosis.

BACKGROUND: Recombinant hirudin, a pure, specific antithrombin could be more effective than heparin in the treatment of deep vein thrombosis, but its short half-life requires constant intravenous infusion, whereas subcutaneous administration of recombinant hirudin can ensure stable and prolonged plasma levels. The aim of our study was to assess the pharmacokinetics, the results on the coagulation variables, and the safety of a recombinant hirudin (HBW 023) administered subcutaneously in patients suffering from deep vein thrombosis. METHODS: Recombinant hirudin (HBW 023) was administered subcutaneously to 10 patients with recent deep vein thrombosis, at a dose of 0.75 mg/kg of body weight twice daily for 5 days, after which standard heparin and acenocoumarol were introduced. Bilateral lower limb venography, and pulmonary angiography, and/or ventilation-perfusion lung scan were carried out on day 1 prior to recombinant hirudin injection and repeated on day 5. aPTT and recombinant hirudin plasma levels were serially assessed after the 1st and the 10th injections. Prothrombin fragments 1 + 2, thrombin-antithrombin III complexes, fibrin degradation products were collected on days 1 and 5. RESULTS: Clinical evolution was uneventful in all but one patient who had a probable recurrence of pulmonary embolism on day 4. No hemorrhagic complication, no untoward biological event was observed. On days 5, Marder score was unchanged or had decreased. Plasma levels of recombinant hirudin peaked in between 3 and 4 h following the injection. aPTT values paralleled, and were significantly correlated with plasma levels of recombinant hirudin on day 1 as well on day 5 (r = 0.903, r = 0.948 respectively). Fragment 1 + 2, and thrombin antithrombin complexes non-significantly decreased from day 1 to day 5. CONCLUSIONS: Subcutaneous administration of recombinant hirudin ensures prolonged stable plasma levels of recombinant hirudin which results in efficient anticoagulation. A dose-ranging study conducted with subcutaneous recombinant hirudin in comparison to conventional heparin therapy may answer the question as to efficacy.

First results obtained in France with the latest model of the Fresenius cell separator: AS 104.

In Besançon, we carried out 40 plateletphereses with the latest model of the Fresenius cell separator AS 104 to check this new system against the new generation of cell separators, according to the following criteria: less than 2x10 6 leukocytes (before filtration) and more than 5x10 11 platelets. The results show that platelet concentrates contained 5.04+/-0.88x10 11 platelets in a total volume of 435+/-113 mL. The mean platelet recovery was 40.95+/-4.86% (from 31.7 to 51.6). The leukocyte content was 2.28+/-5.48x10 6 and the red blood cell contamination was 3.48+/-2.38x10 8. The quality of the platelets was very satisfactory. There was no problem with donor biocompatibility or procedure safety, few adverse donor reactions (0.6%) and good therapeutic efficiency of platelet concentrates.

Changes in plasma fibrin degradation products as a marker of thrombus evolution in patients with deep vein thrombosis.

The level of fibrin degradation products (fdp) as a marker of fibrin clot dissolution was studied prospectively in 51 patients with phlebographically identified deep vein thrombosis (DVT). For this purpose a highly sensitive fdp assay using an anti D neo monoclonal antibody (McAb) was used. At the onset of the hospitalization, in 47 (92%) of the 51 patients tested, the plasma fdp level performed was high, but does not reflect the size of the thrombus, demonstrating that spontaneous thrombus lysis varies from one patient to another. During 10 days of standard heparin or low molecular weight heparin treatment, two different patterns of fdp evolution could be identified in these patients, independent of the type of heparin used. The first was characterized by a gradual decrease in fdp level and a corresponding reduction in the thrombus size. The second pattern showed a persistence of high levels of fdp after 10 days of therapy although the phlebographic score reveals a poor or partial response indicating that fibrinolysis or the balance of thrombus formation/fibrinolysis did not insure total thrombus dissolution. The 4 patients whose initial plasma fdp levels were only slightly increased during the 10 days, seem to have poor thrombolysis, as was shown by the unmodified phlebographic score after 10 days of treatment. Consequently, we conclude that the investigation of plasma fdp levels with a highly sensitive assay should contribute to the evaluation of thrombus evolution in DVT.

[A new treatment of deep venous thrombosis: low molecular weight heparin fractions. Randomized study]. / Un nouveau traitement des thromboses veineuses profondes: les fractions d'héparine de bas poids moléculaire. Etude randomisée.

Sixty-eight patients with acute deep vein thrombosis were allocated at random to two treatment groups. One group (n = 33) received a fixed dose of 750 anti-Xa units of a low molecular weight heparin (CY 222 Choay Institute); the other group (n = 35) received standard heparin in doses of 500 IU/kg/24 h. Both treatments were given for 10 days in two daily subcutaneous injections. A second phlebography was performed on the last day of treatment. No haemorrhagic complication was observed in the group treated with CY 222, as opposed to three cases of haemorrhage in the group treated with standard heparin. The initial phlebographic score and the location of deep vein thrombotic lesions were the same in both groups. Angiographic improvement, with more than 30% thrombolysis, was obtained at the end of treatment in 64% of patients in the CY 222 group and in 65% of patients in the standard heparin group (NS). In 2 patients treated with standard heparin the second phlebography showed extension of the thrombosis. The initial score remained unchanged in 1/3 of patients in both groups. The activated partial thromboplastin time was prolonged (2 or 3 fold the normal value) in the standard heparin group and unchanged in the CY 222 group. Anti-Xa activity was significantly higher in the CY 222 group than in the standard heparin group. It is concluded that CY 222 and standard heparin were equally effective in patients with deep vein thrombosis. However, haemorrhagic complications were more frequent with standard heparin that with CY 222.

[Comparative study of heparin and a very low molecular-weight heparin in hemodialysis in chronic renal insufficiency]. / Etude comparative de l'héparine et d'une héparine de très faible masse moléculaire en hémodialyse dans l'insuffisance rénale chronique.

In the course of dialysis sessions, we have compared the antithrombotic effect of two heparinization regimens: low molecular weight heparin (CY 222, mean molecular weight: 2,500, Institute Choay, France): 90 anti-Xa units/kg bodyweight as a bolus injection followed by a continuous infusion of 1,000 anti-Xa units/hour (regimen 1); or 300 anti-Xa units/kg as a bolus injection (regimen 3), with a standard heparinization regimen (100 IU/kg regimen 2). Eight patients received the 3 regimens successively. Factor IIa and factor Xa inactivation was measured by a method that uses chromogenic substrates. The frequency of adverse effects, ultrafiltration rates, creatinine and BUN clearances of the 3 regimens were similar, whereas dialyser blood loss was higher in the first regimen. At the dose of 300 anti-Xa units of CY 222 (regimen 3), inactivation of factor Xa was similar to Xa inhibition reached through the conventional treatment (regimen 2) but IIa inhibition was less pronounced.

[Efficacy of a very low molecular weight heparin fragment (CY 222) compared to standard heparin in patients with deep venous thrombosis. A randomized study]. / Efficacité d'un fragment d'héparine de trés faible poids moléculaire (CY 222) comparée à l'héparine standard chez des patients porteurs d'une thrombose veineuse profonde. Etude randomisée.

Low molecular weight heparin fractions have been demonstrated as efficient as low doses of standard heparin in preventing post-operative deep-vein thrombosis. Several clinical studies are, now, carrying out in patients with acute deep-vein thrombosis in order to assess efficacy and safety of low molecular weight heparin fractions as compared to standard heparin. In a randomised study including patients with deep-vein thrombosis treated either with low molecular weight heparin or with subcutaneously standard heparin, we demonstrated an effectiveness, in terms of thrombus reduction, of low molecular weight heparin as well as standard heparin. However, low molecular weight heparin seemed to be safer than standard heparin.

Evaluation of two new filtration systems--Fenwal PS400 and Organon Teknika Curesis--and comparison of results with two centrifugation systems--IBM model 2997 and Haemonetics V50.

Two new filtration systems (Fenwal CPS 10TM - PS 400 and Organon Teknika Curesis - M82) were evaluated and compared with two centrifugal cell separators (IBM 2997 and Haemonetics V50). 11 patients with auto immune diseases and dermatological diseases underwent 230 consecutive plasma exchanges. For the filtration systems, the average whole blood rate was 50 ml/min and the plasma separation rate was about 21 ml/min for a transmembrane pressure about 70 mmHg. The pre/post percent reduction and sieving coefficient were calculated for some plasma and blood components. A variety of laboratory studies was monitored to assess the efficacy of plasma separators, their biocompatibility and some yields. These results show that the 2 filters appear safe and efficacious but their modules are too simple and do not offer a great security (no transmembrane pressure control or no extracorporeal fluid balance). For a blood banker, IBM 2997 seems more interesting if we take in account its characteristics during plasma exchanges and the possibility which is offered to carry out cytapheresis procedures. But for a thrombopenic patient the filtration systems keep their advantages.

[Severe thrombopenia from heparin: value of the use of low molecular weight heparin. Apropos of 6 cases]. / Les thrombopénies graves à l'héparine: intérêt de l'utilisation de l'héparine à bas poids moléculaire. A propos de 6 observations.

Severe heparin-induced thrombocytopaenia associated with thromboembolism is a well known complication, although the exact pathogenic mechanism remains unclear. It sets the problem of whether to continue heparin therapy because standard heparin must be withdrawn. Heparin is a mucopolysaccharide composed of fractions of different molecular weights. The fractions with high molecular weights have been held responsible for these severe thrombocytopenias and so, the use of low molecular weight heparin has been suggested. The authors used subcutaneous low molecular weight heparin (CY 216 Choay Institute) at empirical doses of 350 to 1 500 units/kg/24 hour in six cases of severe heparin-induced thrombocytopaenia. Platelet counts rapidly returned to normal (4 days on average) in 5 cases. Thrombocytopaenia persisted with low molecular weight heparin in 1 case. The study of platelet aggregation was positive with low molecular weight heparin in this case and the platelet count returned to normal when the treatment was withdrawn. The authors conclude that, although low molecular weight heparin is useful in severe heparin-induced thrombocytopaenia, its efficacy remains modest. Not only may platelet aggregation persist with low molecular weight heparin which rekindles the debate as to its pathogenic mechanism, but also low molecular weight heparin may have a slight antithrombin effect which limits its use in patients at high risk of thromboembolism, imposing treatment with fast acting vitamin K antagonists.

[Severe thrombopenias during heparin therapy. A phenomenon of intra-vascular platelet aggregation]. / Les thrombopénies majeures au cours d'un traitement héparinique. Phénomène d'agrégation plaquettaire intra-vasculaire.

Haemorrhage or thromboembolism during heparin therapy are usually attributed to a prescribing error. However, these clinical manifestations--especially thromboembolism--may occur with heparin therapy during severe thrombocytopenia. The authors describe the clinical, biological and physiopathological features characterising this thrombocytopenia with reference to 7 personal cases and a review of the literature. The incidence of heparin-induced thrombocytopenia varies between 0.5 and 1%. It seems to be more common (4%) during heparin therapy for thromboembolic disease. The thrombocytopenia appears 8 days after the onset of heparin therapy. It is characterised by the high incidence of thromboembolism (70% of cases) compared to haemorrhagic phenomena (10% of cases). Thrombocytopenia is asymptomatic in 20% of cases. The thrombocytopenia is peripheral, i.e. the bone marrow is normal, and isolated, i.e. there are no deficiencies in the factors of coagulation. One of our cases was of special interest because it was complicated by disseminated intravascular coagulation. Eight cases of disseminated intravascular coagulation have previously been reported. Analysis of platelet aggregation demonstrates the relationship between heparin and thrombocytopenia. Mixing the plasma of patients with thrombocytopenia and plasma rich in platelets in the presence of heparin lead to thrombo-agglutination. In contrast, in control and non-thrombocytopenic heparinised subjects, no reaction was found. These observations prove the existence of a platelet aggregant factor in the plasma during thrombocytopenia. This disappears 6 weeks to 2 months after stopping heparin. This platelet aggregant factor initiates platelet aggregation which is responsible for thrombocytopenia and for the initiation of phenomena of coagulation, so explaining the thromboembolic phenomena.(ABSTRACT TRUNCATED AT 250 WORDS)

[Major thrombopenia induced by heparin treatment. Apropos of 8 cases]. / Les thrombopénies majeures induites par le traitement á l'héparine. A propos de 8 observations.

Major thrombocytopenia occupies a special place among the complications of anticoagulant therapy. Based on a series of 8 personal cases, the authors describe the clinical, biological and physiopathological features of this thrombocytopenia. It occurs in 0.5 to 1 p. 100 of patients on heparin. This figure may not be accurate and thrombocytopenia during heparin therapy occurs almost exclusively in venous thromboembolic disease which it complicated in 4 p. 100 of treated cases. It is detected around the 8th day (between the 4th and 10th day) of treatment and is characterised by thromboembolic events (2 arterial embolisms, 2 pulmonary embolisms, 1 cutaneous necrosis, 1 disseminated intravascular coagulation). Cutaneo-mucous purpura was observed in 1 case. The last two cases in this series were asymptomatic. The thrombocytopenia is peripheral (on average 36,000/mm3) and isolated without any deficiency of the factors of coagulation, except in the case of disseminated intravascular coagulation. A platelet aggregant factor was demonstrated in the plasma of these patients by a thrombo-agglutination technique. This disappeared two months after stopping the heparin. The withdrawal of heparin and relay with oral anticoagulant drugs was associated with a normalisation of the platelet count within 1 to 5 days. Physiopathologically, the high molecular weight fractions of heparin are responsible for this immuno-allergic reaction which leads to the fabrication of a heparin-dependent anti-platelet antibody.