Ibrutinib downregulates a subset of miRNA leading to upregulation of tumor suppressors and inhibition of cell proliferation in chronic lymphocytic leukemia.

The lymph node (LN) is the site of chronic lymphocytic leukemia (CLL) cell activation and proliferation. Aberrant microRNA (miRNA) expression has been shown to have a role in CLL pathogenesis; however, a comparison of miRNA expression between CLL cells in the LN and the peripheral blood (PB) has previously not been reported. On the basis of the analysis of 17 paired LN and PB samples from CLL patients, we identify a panel of miRNAs that are increased in LN CLL cells correlating with an activation phenotype. When evaluated in CLL cells from 38 patients pre and post treatment with ibrutinib, a subset of these miRNAs (miR-22, miR-34a, miR-146b and miR-181b) was significantly decreased in response to ibrutinib. A concomitant increase in putative miRNA target transcripts (ARID1B, ARID2, ATM, CYLD, FOXP1, HDAC1, IBTK, PTEN and SMAD4) was also observed. Functional studies confirmed targets of ibrutinib-responsive miRNAs to include messenger RNA transcripts of multiple tumor suppressors. Knockdown of endogenous miR-34a and miR146b resulted in increased transcription of tumor suppressors and inhibition of cell proliferation. These findings demonstrate that ibrutinib downregulates the expression of a subset of miRNAs related to B-cell activation leading to increased expression of miRNA targets including tumor suppressors and a reduction in cell proliferation.

Association of CD209L tandem repeats polymorphism with susceptibility to human immunodeficiency virus-1 infection, disease progression, and treatment outcomes: a Moroccan cohort study.

In order to investigate the association between length variation of the CD209L neck region and human immunodeficiency virus (HIV)-1 susceptibility, disease progression, and treatment response outcomes, we genotyped 139 HIV-1-seropositive and 109 seronegative individuals. The heterozygous genotype 6/5 showed a significant increased risk of HIV-1 infection (OR 3.03, 95% CI 0.99-9.33, p 0.046). Moreover, after highly active antiretroviral therapy (HAART), HIV-1-seropositive individuals carrying the 6/5, 7/5 and 7/7 genotypes and alleles 5, 6 and 7 showed good CD4(+) T-cell recovery. In addition, individuals with the 7/5, 6/6 and 7/7 genotypes showed a significant decrease in viral load during the treatment period as compared with baseline (p < 0.05). Interestingly, we found that alleles 4 and 6 were associated with protection against AIDS progression. D209L variation may influence susceptibility to HIV-1, response to treatment, and disease progression.

[Rare primary chest wall sarcoma: the synovialosarcoma]. / Sarcome primitif rare de la paroi thoracique: le synovialosarcome.

INTRODUCTION: Malignant primary tumours occurring in the thorax encompass a large group of tumours which may arise from the lung, mediastinal structures, the pleura or the chest wall. OBSERVATION: We report the case of a 37 year old patient, who presented with left sided chest pain. On clinical examination a right sided chest wall mass was identified. Chest X Ray showed a left sided upper mediastinal opacity, associated with a left sided pleural opacity. Thoracic CT scan revealed a large mass arising from the chest wall and infiltrating the mediastinum associated with a second chest wall mass at the level of the 8(th) and 9(th) right ribs. The biopsy of the chest wall mass revealed it to be a parietal synovialosarcoma. The patient responded to chemotherapy based on ifosfamid and doxorubicin as well as mediastino-pulmonary radiotherapy. There was an improvement in the patient's clinical and radiological state but the patient died by pulmonary embolism after the 3(rd) cause of treatment. CONCLUSION: Chest wall synovialosarcoma has a poor prognosis, however, its chemosensitivity means that treatment may initially be effective.

[Enigmatic evolution of an association of pulmonary tuberculosis and amyloidosis]. / Évolution énigmatique d'une association de tuberculose et d'amylose pulmonaire.

INTRODUCTION: Amyloidosis is characterized by tissue deposits of amyloid material. Secondary amyloidosis can occur as a sequel to pulmonary tuberculosis over a relatively long period. However, this was not the case with our patient. Subsequently we conducted a literature review to try to explain the unusual course of AL amyloidosis in our patient. CASE REPORT: A 36- year-old patient was admitted to our department for investigation of haemoptysis. A diagnosis of primary pulmonary tuberculosis was made and antituberculous treatment was started. On the second day of treatment, a haematoma appeared on the sole of the right foot, which spread down to the toes during the following days. Renal investigations showed a 24h proteinuria of 9 g/L and serum protein electrophoresis revealed an albumin level of 11.8 g/L. A diagnosis of nephrotic syndrome was made. A renal biopsy was indicated but this was not possible on account of a marked worsening of the patient's condition after 14 days of treatment. The patient's level of consciousness deteriorated and he was transferred to the intensive care unit for ventilation. He died 48 hours later. Post-mortem histological examination of pulmonary and cutaneous tissue revealed AL amyloid deposits. CONCLUSION: In view of the association of active pulmonary tuberculosis and a pulmonary localisation of amyloidosis, a causal relationship is not definite. Coexistence of active pulmonary tuberculosis and primary amyloidosis must also be considered, particularly as the immunohistochemical characterisation revealed AL amyloidosis.

[Neuropsychiatric side effects of antituberculosis agents]. / Les effets indésirables neuropsychiatriques des antituberculeux.

UNLABELLED: Accurency of neurological or psychiatric complications secondary to the administration of antituberculosis may be at the origin of diagnosis and therapeutic problems. This work aims at studying the frequency of these manifestations, their clinical presentations and their therapeutic approach. PATIENTS AND METHODS: This retrospective study was carried on from January 1990 to June 2008 at Ibn Nafis Pulmonary Department of Abderrahmen Mami Hospital in Ariana. It dealt with 18 in-patients with a neurologic or a psychiatric complication due to antituberculous drugs. Peripheral neuropathy was noted in 6 patients (33%). One of them had a history of chronic alcoolism, another one had a history of diabetes mellitus and 2 other patients were more than 72 years old. We had definitively stopped isoniazide in 2 cases and decreased the dosage in 4 other patients. However, all patients received B6 vitamin. Convulsions occured in 2 women without any history of epileptic status. Anticonvulsivant treatment was prescribed and isoniazid definitively stopped. Hallucinosis was noted in 4 patients, with one having a history of chronic alcoolism. Isoniazide was stopped in all cases. Agressivity, insomnia and memory problems were noted in 6 patients. Isoniazid was interrupted in only one woman who had history of depression. In 5 other patients, anxiolytics were prescribed. Isoniazide was incrimined in all cases and evolution was favorable for the 18 patients. A close monitoring of patients on antituberculous treatment is required to detect the onset of any neuropsychiatric complications incriminating usually isoniazid. Definitive interruption or decrease of the dose of isoniazid depending of the acetylation test were necessary.

[Hypersensitivity reactions to antituberculous therapy]. / Manifestations d'hypersensibilité aux antituberculeux.

INTRODUCTION: Hypersensitivity reactions to antituberculous drugs pose both a diagnostic and a therapeutic problem. This work aims to study the frequency of allergic reactions to antituberculous drugs, their clinical presentations and the diagnostic approach required. PATIENTS AND METHODS: This retrospective study covered the period from January 1990 to June 2008 at the Ibn Nafis Pulmonary Department of Abderrahmen Mami Hospital in Ariana. It dealt with 30 in-patients who experienced an allergic reaction to antituberculous drugs. RESULTS: Cutaneous manifestations were the most frequent (80% of cases), predominantly urticarial. Thrombocytopenia was noted in two cases, anaphylactic shock in three cases, a systemic toxidermia in two cases and renal failure in one patient. Pyrazinamide was implicated in most cases (28%) when only one drug was considered to be responsible for the reaction. Interrupting either one drug or the whole treatment was necessary to define the cause of the reaction. The clinical evolution of hypersensitivity signs was favorable in all cases following definitive withdrawal of the responsible drug. Complete recovery from tuberculosis occurred in all cases. CONCLUSION: Close monitoring of patients on antituberculous treatment is required to detect the onset of any allergic reaction and ensure an adequate compliance with treatment.

[Effect of tobacco smoking on pulmonary tuberculosis]. / Effet du tabagisme sur la tuberculose pulmonaire.

Tobacco increases the risk of pulmonary infection, especially tuberculosis. We try by this study to analyse this action. It's a comparative study between two groups of patients hospitalized in our department between January 2006 and June 2008. The first group was made of 30 smokers patients hospitalized because of confirmed pulmonary tuberculosis. The second group consists of 30 non smokers patients and also hospitalized because of confirmed pulmonary tuberculosis. Delay of diagnosis was longer in the group of smokers (3, 1 +/- 3 months versus 2 +/- 1 month with p = 0.039). The most frequent symptoms in both groups were cough and loss of weight. The biological investigations showed a high level of white blood cells with predominance of neutrophil cells especially in smokers (p = 0.024). Chest X ray showed essentially nodules in both groups. These nodules were bilateral in smokers (P = 0.045). Evolution after antituberculosis treatment was favourable for all patients. A delay of recovery (time between symptoms and recovery) was longer in smokers than in non smoker patients (p = 0.043). Pulmonary Sequels such as dyspnoea (p = 0.016) and fibrosis (p = 0.041) were most frequent in smokers. No patients had tuberculosis relapse. Tobacco may delay the recovery of pulmonary tuberculosis and may induce pulmonary sequels in spite of correctly antituberculosis treatment.

Multiple myeloma presenting with multiple thoracic manifestations.

Multiple myeloma is a malignant proliferation of plasma cells that affects mainly bone marrow but may also involve other organs as well. We report thoracic involvement in the form of left-sided pleural effusion, osseous lesions, bronchial infiltration, and mediastinal lymphadenopathy in a 61-year-old woman, non-smoker presented with chest pain, dyspnoea, cough and deterioration in general health over the preceding seven months. Immunoelectrophoresis and immunofixation showed raised kappa-light chain immunoglobulin G (IgG) in serum and pleural fluid. Bronchial and pleural biopsies documented myelomatous infiltration and bone marrow aspirate revealed extensive plasma cell infiltration. At eight months, following the fourth cycle of melphalan, endoxan and prednisone based chemotherapy, the patient died.

Aided air conduction threshold as a predictor of cochlear reserve in otosclerosis patients.

Otosclerosis classically presents with conductive deafness. However, mainly in older patients or those with a progressive (active) form of this disease, it presents with elevated bone conduction (BC) thresholds. In many cases, an elevation in the BC threshold during otosclerosis is not a true indicator of 'cochlear reserve' and this apparent loss can be corrected by surgery. It is generally accepted that in those patients, elevated BC thresholds are not due to pure sensorineural hearing loss but to the Carhart effect. Speech discrimination tests are routinely done, and the results of these tests determine if a patient is a good candidate for surgery. In most institutions, those with poor speech discrimination scores are considered to be poor candidates due to inadequate cochlear reserve. Using hearing-aid-assisted pure-tone audiometry, we developed an accurate method to identify suitable candidates for surgery among the patients with elevated BC thresholds. This method is meant to complement speech discrimination tests in predicting the success of surgery in those patients. Different variables render speech discrimination tests unreliable in many patients. The rationale behind the current study was the lack of an easy and accurate clinical method to determine if a patient has a good cochlear reserve, and to predict the postoperative hearing threshold outcome.

[Pleural empyema revealing osteopoikilosis]. / Pleurésie purulente révélatrice d'une ostéopoecilie.

INTRODUCTION: Osteopoikilosis is a rare, inherited and usually asymptomatic sclerosing bone dysplasia of unknown etiology which predominantly involves the appendicular and rarely the axial skeleton. CASE REPORT: We report the case of a 24 year old man who was hospitalized for pleural empyema and treated with antibiotics for six weeks in addition to pleural evacuation and physiotherapy. The diagnosis of osteopoikilosis associated with the pleural empyema was made on the radiological findings. In fact the chest X-Ray showed spherical areas of increased bone density in both humeral epiphyses. In order to explore these bone abnormalities further investigations were performed, including red and white blood cell counts, sedimentation rate and protein electrophoresis. There were no biological abnormalities. Radiography of the whole skeleton showed disseminated sclerotic lesions in the pelvis and the metacarpal and carpal bones of both hands. A neoplastic aetiology was excluded. In the light of these investigations, the diagnosis of osteopoikilosis was established. CONCLUSION: Widespread osteopoikilosis can be revealed on chest radiography.

Interaction of macrolides and ketolides with the phagocytic cell line PLB-985.

Interactions between antibacterial agents and polymorphonuclear neutrophils (PMNs) are a major focus of investigation. Owing to the variable drug susceptibility of PMNs from different individuals, in vitro studies require samples from large panels of healthy volunteers to reach statistical significance. Here, we used a phagocytic cell line, PLB-985, which can differentiate into mature PMNs in vitro, for the study of cellular interactions (drug uptake and antioxidant effects) of two macrolides (azithromycin and roxithromycin) and four ketolides [HMR 3004, HMR 3647 (telithromycin), HMR 3562 and HMR 3787]. The oxidative burst of differentiated (D) cells was inhibited by macrolides and ketolides. IC50% values (concentrations impairing the oxidative burst by 50%), determined after 30 min of incubation, were as follows for azithromycin, roxithromycin, HMR 3004, telithromycin, HMR 3562 and HMR 3787, respectively: 40, 39, 15, 23, 26, and 33 mg/l (fMLP stimulation) and 37, 86, 39, 43, 14, and 31 mg/l (PMA stimulation). These values were similar to those obtained with PMNs. Uptake of the two macrolides was significantly lower in non-differentiated (ND) cells than in D cells and PMNs. The cellular/extracellular (C/E) concentration ratios at 60 min for PMNs, D and ND PLB were respectively 67, 25 and 11 (roxithromycin) and 159, 137 and 48 (azithromycin). Ketolide uptake by ND-PLB was also significantly lower than that obtained with PMNs (C/E ratios at 60 min were about 75 versus 265 (HMR 3004), 36 vs 230 (telithromycin), 75 vs 235 (HMR 3562) and 20 vs 130 (HMR 3787). Although the active carrier system seemed to be present in ND cells, its activation pathway was not functional. Thus, the PLB-985 cell line is a good in vitro model for studying drug-PMN interactions. The use of ND and D cells may shed light on the nature and activation pathways of macrolide transport systems present on the PMN membrane.

Structure-activity relationships among 9-N-alkyl derivatives of erythromycylamine and their effect on the oxidative burst of human neutrophils in vitro.

Macrolide antibiotics have recently triggered much interest owing to the immunomodulatory potential of some derivatives, particularly in the field of inflammatory diseases. Among the possible mechanisms underlying these anti-inflammatory effects, macrolide-induced inhibition of oxidant production by phagocytes has attracted much attention. We and others have previously reported that erythromycin A-derived macrolides impair the phagocyte oxidative burst, a property linked to the presence of L-cladinose. However, we have also demonstrated that other substituents can be involved in the modulation of phagocyte function. Here we have extended the analysis of structure-activity relationships by studying the effects of five 9-N-alkyl derivatives of erythromycylamine on oxidant production by human neutrophils in vitro. LY211397 (2-methoxyethyl derivative) neither altered cell viability nor superoxide anion production. LY281389 (n-propyl derivative) did not alter cell viability and was slightly more inhibitory than erythromycylamine for the production of superoxide anion; its IC50 (concentration that inhibits 50% of the neutrophil response) was about 18 and 24 microM (versus 72 and 74 pM for erythromycylamine) after 60 min of incubation following fMLP and PMA stimulation, respectively. LY80576 (N-phenyl-3-indolylmethyl derivative), LY281981 (3-phenyl-n-propyl derivative) and LY57843 (benzyl derivative) all displayed cellular toxicity at high pharmacological concentrations after 30 to 60 min of incubation. Interestingly, these latter three drugs exhibited a rapid (5 min incubation) and strong inhibitory effect on the neutrophil oxidative burst from either stimulus, with IC50 values of 3 to 10 pM. Further in-vitro and in-vivo investigations are required to analyze the anti-inflammatory potential of these three derivatives.

Effect of telithromycin (HMR 3647) on polymorphonuclear neutrophil killing of Staphylococcus aureus in comparison with roxithromycin.

HMR 3647 (telithromycin), a new ketolide, is active on intracellular pathogens. It was previously demonstrated that it inhibits superoxide anion production in a time- and concentration-dependent manner, at concentrations which inhibit 50% of the control response of about 55 microg/ml (5 min) to 30 microg/ml (30 min); these values are similar to those obtained with roxithromycin, a classical erythromycin A derivative. Here we investigated whether these drugs modified the bactericidal activity of human polymorphonuclear neutrophils (PMN) on four strains of Staphylococcus aureus with different profiles of susceptibility to macrolides and ketolides. We found that the main factor involved in killing was the antibacterial potency of the drugs, although combinations of antibiotics with PMN were slightly more active than each component used alone against two of the four strains. In addition, high concentrations of the drugs, which impaired the PMN oxidative burst, did not impair PMN bactericidal activity. Likewise, some cytokines which enhance PMN oxidative metabolism did not modify PMN bactericidal activity in the presence or absence of macrolides or ketolides. These data suggest that oxygen-independent mechanisms contribute to the bactericidal activity of PMN on these strains of S. aureus. Both live and/or heat-killed bacteria impaired the uptake of telithromycin and roxithromycin (but not that of levofloxacin, a quinolone) in a concentration-dependent manner, owing to a modulation of PMN transductional systems involved in the activation of the macrolide carrier.

Cellular uptake of two fluoroketolides, HMR 3562 and HMR 3787, by human polymorphonuclear neutrophils in vitro.

We analyzed the cellular accumulation of two new fluoroketolides, HMR 3562 and HMR 3787, by human polymorphonuclear neutrophils (PMN) in vitro. Both compounds were rapidly taken up by PMN, with a cellular-to-extracellular concentration ratio (C/E) of about 141 (HMR 3562) and 117 (HMR 3787) at 5 min, and this was followed by a plateau at 60 to 180 min, with a C/E of >300 at 180 min. Both ketolides were mainly located in PMN granules (about 75%) and egressed slowly from loaded cells (about 40% at 60 min), owing to avid reuptake. Uptake was moderately sensitive to external pH, and activation energy was also moderate (about 70 kJ/mol). As with other macrolides and ketolides, the existence of an active transport system was suggested by (i) the strong interindividual variability in uptake kinetics, suggesting variability in the number or activity of a transport protein; (ii) the saturation kinetics characteristic of a carrier-mediated transport system (V(max), about 2,300 ng/2.5 x 10(6) PMN/5 min; K(m), about 50 microg/ml); (iii) the inhibitory effects of Ni(2+) (a blocker of the Na+-Ca(2+) exchanger), phorbol myristate acetate (a protein kinase C activator), and H89 (a protein kinase A inhibitor). Although these two ketolides are more related to HMR 3647 (telithromycin), it is interesting that the presence of a fluoride gave these molecules a cellular pharmacokinetics more like those of HMR 3004 than those of HMR 3647. The macrolide transport system has not been yet elucidated, but our data confirm that, despite variations in chemical structure, all erythromycin A derivatives share a transmembrane transport system.

Immunomodulation by macrolide antibiotics.

Macrolide antibiotics are strongly concentrated within host cells, a property that sustains their activity against intracellular pathogens and is likely responsible for the modulation of cell metabolism and function. There is extensive literature on the subject of macrolide-induced modulation of immune responses. Erythromycin A derivatives seem to display anti-inflammatory activity in vitro, in some animal models and in various clinical settings such as diffuse panbronchiolitis (DPB). The underlying mechanisms are not yet fully understood: inflammatory cytokine and oxidant production by phagocytes is down-regulated by these drugs, but other possible targets include bacterial virulence factors, bronchial and epithelial cells, etc. Also, a link has been suggested between the macrolide transmembrane carrier system and the P-glycoprotein family, which comprises MDR (multiple drug resistance) and CFTR (cystic fibrosis transmembrane conductance regulator), which are respectively involved in the chemotherapeutic resistance of cancer cells and in the genesis of cystic fibrosis.

Erythromycin A-derived macrolides modify the functional activities of human neutrophils by altering the phospholipase D-phosphatidate phosphohydrolase transduction pathway: L-cladinose is involved both in alterations of neutrophil functions and modulation of this transductional pathway.

All erythromycin A derivatives, irrespective of the size of the lactone ring and the nature of the substituent, inhibit oxidant production by neutrophils and promote their degranulation. We demonstrate in this study that the L-cladinose at position 3 of the lactone ring is a key structure in the modulation of these two neutrophil functions, suggesting that this sugar (alone or combined with a lactone structure) interferes with cell target(s) involved in both oxidant production and exocytosis. Taking roxithromycin as an example of erythromycin A derivatives, we also found that these molecules interfered with the phospholipase D (PLD)-phosphatidate phosphohydrolase pathway in two ways. In nonstimulated neutrophils, roxithromycin and all L-cladinose-bearing molecules activated PLD, as reflected by 1-O-[3H]alkyl-2-acyl-phosphatidyl-ethanol production. In addition, these drugs induced an accumulation of 1-O-[3H]alkyl-2-acyl-phosphatidic acid (PA), but not 1-O-[3H]alkyl-2-acylglycerol. PA accumulation seems to be involved in the induction of exocytosis by macrolides, as the roxithromycin-induced release of granular enzymes was impaired strongly in the presence of ethanol. By contrast, in stimulated neutrophils, roxithromycin inhibited PLD activity and totally impaired 1-O-[3H]alkyl-2-acylglycerol production. The inhibition of diglyceride production by roxithromycin (not its descladinosyl derivative) could explain its inhibitory effect on oxidant production. The relevance of our data to the clinical situation, particularly the anti-inflammatory activity of these drugs, requires further investigation.

Cellular accumulation of the new ketolide RU 64004 by human neutrophils: comparison with that of azithromycin and roxithromycin.

We analyzed the uptake of RU 64004 by human neutrophils (polymorphonuclear leukocytes [PMNs]) relative to those of azithromycin and roxithromycin. RU 64004 was strongly and rapidly accumulated by PMNs, with a cellular concentration/extracellular concentration ratio (C/E) of greater than 200 in the first 5 min, and this was followed by a plateau at 120 to 180 min, with a C/E of 461 +/- 14.8 (10 experiments) at 180 min. RU 64004 uptake was moderately sensitive to external pH, and activation energy was also moderate (63 +/- 3.8 kJ/mol). RU 64004 was mainly located in PMN granules (about 70%) and egressed slowly from loaded cells, owing to avid reuptake. The possibility that PMN uptake of RU 64004 and other macrolides occurs through a carrier-mediated system was suggested by three key results. First, there existed a strong interindividual variability in uptake kinetics, suggesting variability in the numbers or activity of a transport protein. Second, macrolide uptake displayed saturation kinetics characteristic of that of a carrier-mediated transport system: RU 64004 had the highest Vmax value (3,846 ng/2.5 x 10(6) PMNs/5 min) and the lowest Km value (about 28 microM), indicating a high affinity for the transporter. Third, as observed previously with other erythromycin A derivatives, Ni2+ (a blocker of the Na+/Ca2+ exchanger which mediates Ca2+ influx in resting neutrophils) impaired RU 64004 uptake by PMNs, with a 50% inhibitory concentration of about 3.5 mM. In addition, we found that an active process is also involved in macrolide efflux, because verapamil significantly potentiated the release of all three macrolides tested. This effect of verapamil does not seem to be related to an inhibition of Ca2+ influx, because neither EGTA [ethylene glycol-bis (beta-aminoethyl ether)-N,N',N'-tetraacetic acid] nor Ni2+ modified macrolide efflux. The nature and characteristics of the entry- and efflux-mediating carrier systems are under investigation.

Comparison of various macrolides on stimulation of human neutrophil degranulation in vitro.

Macrolide antibiotics are taken up and concentrated by host cells, particularly phagocytes, and are likely candidates to modify cell functions. In this study, we extended our previous work concerning the effect of three 14-membered-ring macrolides (dirithromycin, erythromycin and erythromycylamine) on human neutrophil exocytosis, and found that three other erythromycin A derivatives (roxithromycin, clarithromycin and the azalide, azithromycin) also triggered neutrophil degranulation in a time- and concentration-dependent manner. After 30 min of incubation, the correlation coefficients for concentration-dependence for roxithromycin were 0.885, 0.739 and 0.750 (P < 0.005) and for clarithromycin were 0.795, 0.599, 0.733 (P < 0.02), respectively, for lysozyme, beta-glucuronidase and lactoferrin release. Although the underlying mechanism was not elucidated, these and previous data suggest that intracellular accumulation is a prerequisite. Furthermore, comparison of the characteristics of macrolide-induced exocytosis with those of exocytosis triggered by the synthetic chemotactic stimulus FMLP suggested that different mechanisms are involved. In keeping with this possibility, we showed that combined treatment (macrolides plus FMLP) resulted in totally additive exocytosis of azurophilic but not specific granules. The clinical relevance of our data remains to be ascertained.