Laurent Guilleminault

Noninvasive diagnosis of actionable mutations by deep sequencing of circulating free DNA in lung cancer from never-smokers: a proof-of-concept study from BioCAST/IFCT-1002.

Purpose: Tumor somatic mutation analysis is part of the standard management of metastatic lung cancer. However, physicians often have to deal with small biopsies and consequently with challenging mutation testing. Circulating free DNA (cfDNA) is a promising tool for accessing the tumor genome as a liquid biopsy. Here, we evaluated next-generation sequencing (NGS) on cfDNA samples obtained from a consecutive series of patients for the screening of a range of clinically relevant mutations. Experimental Design: A total of 107 plasma samples were collected from the BioCAST/IFCT-1002 lung cancer study (never-smokers cohort). Matched tumor DNA (tDNA) was obtained for 68 cases. Multiplex PCR-based assays were designed to target specific coding regions in EGFR, KRAS, BRAF, ERBB2, and PI3KCA genes, and amplicon sequencing was performed at deep coverage on the cfDNA/tDNA pairs using the NGS IonTorrent Personal Genome Machine Platform. Results: CfDNA concentration in plasma was significantly associated with both stage and number of metastatic sites. In tDNA, 50 mutations (36 EGFR, 5 ERBB2, 4 KRAS, 3 BRAF, and 2 PIK3CA) were identified, of which 26 were detected in cfDNA. Sensitivity of the test was 58% (95% confidence interval, 43%–71%) and the estimated specificity was 87% (62%–96%). Conclusion: These data demonstrate the feasibility and potential utility of mutation screening in cfDNA using IonTorrent NGS for the detection of a range of tumor biomarkers in patients with metastatic lung cancer. Clin Cancer Res; 20(17); 4613–24. ©2014 AACR.

Fate of inhaled monoclonal antibodies after the deposition of aerosolized particles in the respiratory system.

Monoclonal antibodies (mAbs) are usually delivered systemically, but only a small proportion of the drug reaches the lung after intravenous injection. The inhalation route is an attractive alternative for the local delivery of mAbs to treat lung diseases, potentially improving tissue concentration and exposure to the drug while limiting passage into the bloodstream and adverse effects. Several studies have shown that the delivery of mAbs or mAb-derived biopharmaceuticals via the airways is feasible and efficient, but little is known about the fate of inhaled mAbs after the deposition of aerosolized particles in the respiratory system. We used cetuximab, an anti-EGFR antibody, as our study model and showed that, after its delivery via the airways, this mAb accumulated rapidly in normal and cancerous tissues in the lung, at concentrations twice those achieved after intravenous delivery, for early time points. The spatial distribution of cetuximab within the tumor was heterogeneous, as reported after i.v. injection. Pharmacokinetic (PK) analyses were carried out in both mice and macaques and showed aerosolized cetuximab bioavailability to be lower and elimination times shorter in macaques than in mice. Using transgenic mice, we showed that FcRn, a key receptor involved in mAb distribution and PK, was likely to make a greater contribution to cetuximab recycling than to the transcytosis of this mAb in the airways. Our results indicate that the inhalation route is potentially useful for the treatment of both acute and chronic lung diseases, to boost and ensure the sustained accumulation of mAbs within the lungs, while limiting their passage into the bloodstream.

Emergence of community-acquired Clostridium difficile infection: the experience of a French hospital and review of the literature

BACKGROUND Clostridium difficile infection (CDI) is a common cause of nosocomial diarrhoea. People in the general community are not usually considered to be at risk of CDI. CDI is associated with a high risk of morbidity and mortality. The risk of severity is defined by the Clostridium Severity Index (CSI). METHODS The cases of 136 adult patients with CDI treated at the University Hospital of Tours, France between 2008 and 2012 are described. This was a retrospective study. RESULTS Among the 136 patients included, 62 were men and 74 were women. Their median age was 64.4 years (range 18-97 years). Twenty-six of the 136 (19%) cases were community-acquired (CA) and 110 (81%) were healthcare-acquired (HCA). The major risk factors for both groups were long-term treatment with proton pump inhibitors (54% of CA, 53% of HCA patients) and antibiotic treatment within the 2.5 months preceding the CDI (50% of CA, 91% of HCA). The CSI was higher in the CA-CDI group (1.56) than in the HCA-CDI group (1.39). Intensive care was required for 8% of CA-CDI and 16.5% of HCA-CDI patients. CONCLUSIONS CDI can cause community-acquired diarrhoea, and CA-CDI may be more severe than HCA-CDI. Prospective studies of CDI involving people from the general community without risk factors are required to confirm this observation.

VEGF neutralizing aerosol therapy in primary pulmonary adenocarcinoma with K-ras activating-mutations

K-ras mutations promote angiogenesis in lung cancer and contribute to the drug resistance of cancer cells. It is not clear whether K-ras mutated adenocarcinomas are sensitive to anti-angiogenic therapy with monoclonal antibodies (mAbs) that target vascular endothelial growth factor (VEGF). Anti-angiogenic mAbs are usually delivered systemically, but only a small proportion reaches the lung after intravenous injection. We investigated the relevance of a non-invasive pulmonary route for the delivery of anti-VEGF mAbs in the mouse K-rasLA1 model. We found that pulmonary delivery of these mAbs significantly reduced the number of tumor lesions and inhibited malignant progression. The antitumor effect involves the VEGFR2-dependent inhibition of blood vessel growth, which impairs tumor proliferation. Pharmacokinetic analysis of aerosolized anti-VEGF showed its low rate of passage into the bloodstream, suggesting that this delivery route is associated with reduced systemic side effects. Our findings highlight the value of the aerosol route for administration of anti-angiogenic mAbs in pulmonary adenocarcinoma with K-ras activating-mutations.

Asthma Unmasked With Tumor Necrosis Factor-α-Blocking Drugs

We report five cases of asthma unmasked by anti-tumor necrosis factor (TNF)-α-blocking drugs. Asthma symptoms appeared within an average of 4 months (range 1-24 months) after starting the anti-TNF-α treatment for inflammatory disease. The patients did not appear to be predisposed to asthma except for one patient who had asthma during childhood. Four patients stopped anti-TNF-α-blocking drugs with an improvement of symptoms within 1 to 5 months. In the patient with a history of childhood asthma, respiratory symptoms recurred when another anti-TNF-α therapy was started. Asthma control was achieved with inhaled steroids, allowing anti-TNF-α treatment to continue. The biotherapy was maintained for the fifth patient in association with inhaled steroids. The pathophysiologic mechanisms are unknown but are probably more complex than the T helper 1/T helper 2 imbalance suggested in the literature, and further studies are required.

Gastroesophageal reflux disease is a risk factor for severity of organizing pneumonia.

Background: The link between organizing pneumonia (OP) and gastroesophageal reflux disease (GERD) is not well known. There is little evidence in the literature to establish a causal link between GERD and OP. Objectives: The aim of the study was to assess the hypothesis that OP is more severe when it is associated with GERD and that it leads to more frequent relapses. Methods: In a retrospective study on 44 patients suffering from OP, we compared the clinical, radiological and histological characteristics of 2 groups, 1 composed of patients with GERD (n = 20) and the other of patients without GERD (n = 24). Results: The GERD group was distinguished by a higher number of patients with migratory alveolar opacities on chest radiography and thoracic computerized tomography (14/20 vs. 9/24; p = 0.03 and 18/20 vs. 13/24; p = 0.01), greater hypoxemia [60 (42-80) vs. 70 (51-112) mm Hg; p = 0.03], greater bronchoalveolar lavage cellularity [0.255 (0.1-1.8) vs. 0.150 (0.05-0.4) g/l; p = 0.035] and more frequent relapses (14/20 vs. 9/24; p = 0.03). Conclusions: OP associated with GERD is more severe and results in more frequent relapses. Microinhalation of gastric secretions might induce lung inflammation leading to OP and relapse. We suggest that typical symptoms of GERD such as pyrosis should be investigated in OP.

Diffuse panbronchiolitis and IgA nephropathy

Chronic infections of respiratory tracts are suspected to promote the development of IgA nephropathy. From the study of one case, we discuss the hypothesis that infections by pseudomonas aeruginosa might promote an increased bronchial production of IgA1, which could pass in the serum and settle into immune complexes in the kidney. A 67 years old woman presented simultaneously IgA nephropathy and diffuse panbronchiolitis. The evolution was marked by repeated infections by pseudomonas aeruginosa. The study of lung biopsy by immunohistochemistry showed intense expression of IgA1 in bronchioles, increased when compared to large bronchi. In contrast, expression of the transport receptor (pIgR) was decreased in inflamed bronchioles and preserved in bronchi. The BAL during an infection by pseudomonas aeruginosa showed increased secretory (S-) IgA with predominance of S-IgA1 (28.6 µg/mL versus S-IgA2 8.4µg/mL). In the sera collected during two infectious episodes and compared with an inter-critical sample, we found an increased IgA1 (776 and 549µg/mL versus 455µg/mL), associated with increased polymeric IgA (estimated at 40-50%, versus normally  10%). The increased expression of IgA1 in bronchiolar tissue, BAL and serum in the case of our patient further suggests a putative link between IgA nephropathy and diffuse panbronchiolitis, through exuberant production of IgA1 induced by pseudomonas aeruginosa infections.

Aerosoltherapy in lung cancer

Introduction: Although airways route is an attractive alternative route to systemic administration, increasing concentration of the drug in the lungs while reducing whole-body toxicity, it is not often exploited in lung cancer treatment. After checking that cetuximab, an anti-cancer antibody binding to EGFR, resists the physical constraints of nebulization, we established an animal model of lung tumour sensitive to the antibody. Then, we used the animal model to evaluate cetuximab antitumor efficiency, biodistribution and pharmacokinetic following delivery through airways. Methods: Cetuximab was nebulized with the IA-1C MicroSprayer™ (PennCentury Inc., USA) connected to a FMJ-250 high pressure syringe, a device used to administrate, directly inside the trachea, aerosol in mice. The effect of nebulization on cetuximab was assessed in terms of its affinity for membrane EGFR (using flow cytometry), inhibition of cell growth and inhibition of EGFR phosphorylation. To ensure that airways delivery of cetuximab did not lead to additional toxicity, we compared tolerance to cetuximab delivered through systemic and pulmonary routes in mice without tumors. An animal model of lung cancer sensitive to cetuximab was established and consists in the instillation of human epidermoid carcinoma cells endotracheally in nude mice. The model was exploited to study cetuximab biodistribution and pharmacokinetic following delivery through systemic and pulmonary routes. Therapeutic efficiency of nebulized cetuximab was determined in tumor- bearing animals. Results: Firstly, our results showed that MicroSprayer™ did not alter cetuximab integrity, immunological and pharmacological properties. Secondly, airways administration of cetuximab in mice seemed to be well-tolerated and did not induced additional toxicity in lungs, kidney, colon, skin, liver or spleen. Finally, higher, more rapid and prolonged lung uptake was observed in animals receiving cetuximab through the pulmonary route. Antitumor efficiency of nebulized cetuximab is currently under investigation. Conclusion: These results highlight the potential of the pulmonary route for delivery of anticancer antibody in lung cancer.