Excessive bone formation in a mouse model of ankylosing spondylitis is associated with decreases in Wnt pathway inhibitors

Introduction: Ankylosing spondylitis (AS) is unique in its pathology where inflammation commences at the entheses before progressing to an osteoproliferative phenotype generating excessive bone formation that can result in joint fusion. The underlying mechanisms of this progression are poorly understood. Recent work has suggested that changes in Wnt signalling, a key bone regulatory pathway, may contribute to joint ankylosis in AS. Using the proteoglycan-induced spondylitis (PGISp) mouse model which displays spondylitis and eventual joint fusion following an initial inflammatory stimulus, we have characterised the structural and molecular changes that underlie disease progression. Methods: PGISp mice were characterised 12 weeks after initiation of inflammation using histology, immunohistochemistry (IHC) and expression profiling. Results: Inflammation initiated at the periphery of the intervertebral discs progressing to disc destruction followed by massively excessive cartilage and bone matrix formation, as demonstrated by toluidine blue staining and IHC for collagen type I and osteocalcin, leading to syndesmophyte formation. Expression levels of DKK1 and SOST, Wnt signalling inhibitors highly expressed in joints, were reduced by 49% and 63% respectively in the spine PGISp compared with control mice (P < 0.05) with SOST inhibition confirmed by IHC. Microarray profiling showed genes involved in inflammation and immune-regulation were altered. Further, a number of genes specifically involved in bone regulation including other members of the Wnt pathway were also dysregulated. Conclusions: This study implicates the Wnt pathway as a likely mediator of the mechanism by which inflammation induces bony ankylosis in spondyloarthritis, raising the potential that therapies targeting this pathway may be effective in preventing this process.

Genetic studies of chondrocalcinosis

Purpose of review Our understanding of the causation of the chondrocalcinosis and other disorders characterized by ectopic mineralization is rapidly increasing, and genetic studies have contributed substantially to recent major advances in the field. This review will discuss what is known about the genetics of chondrocalcinosis and what we have learned from genetic studies to date. Recent findings: Chondrocalcinosis is one of a family of conditions associated with ectopic mineralization. This family also includes disorders of mineralization of bone and spinal and other ligaments, and vascular calcification. There has been increasing evidence of the key role of transport and metabolism of inorganic pyrophosphate (PPi) in control of mineralization, and as the likely explanation for the association of a variety of genetic variants with chondrocalcinosis and ectopic mineralization elsewhere. This may be an overly simplistic view of this family of conditions, with recent evidence suggesting that, for example, ANKH variants may not all predispose to chondrocalcinosis by effects on PPi transport, but may also influence chondrocyte maturation. Summary: Understanding the control of the process of mineralization and its tissue specificity are important steps in the search for rational therapies for these conditions.

PREPARE Study: Patient satisfaction survey with care provided in the low clearance clinic

Background Formalised predialysis care has been shown to extend the wellness of individuals with advanced chronic kidney disease, slow disease progression and increase the uptake of home dialysis. Predialysis care, incorporating multidisciplinary input is also vital in delaying the onset of end-stage kidney disease and reducing hospital admissions; thereby decreasing financial demands on health budgets. Predialysis care should include comprehensive information provision and predialysis education. This empowers patients to choose self-care strategies and therapies.

Consensus statement on the investigation and management of non-radiographic axial spondyloarthritis (nr-axSpA)

Aim Non-radiographic axial spondyloarthritis (nr-axSpA) is axial inflammatory arthritis where plain radiographic damage is not evident. An unknown proportion of these patients will progress to ankylosing spondylitis (AS). The increasing recognition of nr-axSpA has been greatly assisted by the widespread use of magnetic resonance imaging. The aim of this article was to construct a set of consensus statements based on a literature review to guide investigation and promote best management of nr-axSpA. Methods A literature review using Medline was conducted covering the major investigation modalities and treatment options available. A group of rheumatologists and a radiologist with expertise in investigation and management of SpA reviewed the literature and formulated a set of consensus statements. The Grade system encompassing the level of evidence and strength of recommendation was used. The opinion of a patient with nr-axSpA and a nurse experienced in the care of SpA patients was also sought and included. Results The literature review found few studies specifically addressing nr-axSpA, or if these patients were included, their results were often not separately reported. Fourteen consensus statements covering investigation and management of nr-axSpA were formulated. The level of agreement was high and ranged from 8.1 to 9.8. Treatment recommendations vary little with established AS, but this is primarily due to the lack of available evidence on the specific treatment of nr-axSpA. Conclusion The consensus statements aim to improve the diagnosis and management of nr-axSpA. We aim to raise awareness of this condition by the public and doctors and promote appropriate investigation and management.

Early detection of melanoma metastases using microRNAs as novel biomarkers

Early detection of melanoma skin cancer, prior to metastatic spread, is critical to improve survival outcomes in patients. This study identified a melanoma-related panel of blood markers that can detect the presence of melanoma with high sensitivity and accuracy which is superior to currently used markers for melanoma progression, recurrence, and survival. Overall, the findings discussed in this thesis may lead to more precise measurement of disease progression allowing for better treatments and an increase in overall survival.

Analysis of a Genomic DNA Expression Library of Mycobacterium tuberculosis Using Tuberculosis Patient Sera: Evidence for Modulation of Host Immune Response

DNA obtained from a human sputum isolate of Mycobacterium tuberculosis, NTI-64719, which showed extensive dissemination in the guinea pig model resulting in a high score for virulence was used to construct an expression library in the lambda ZAP vector. The size of DNA inserts in the library ranged from 1 to 3 kb, and recombinants represented 60% of the total plaques obtained. When probed with pooled serum from chronically infected tuberculosis patients, the library yielded 176 recombinants with a range of signal intensities. Among these, 93 recombinants were classified into 12 groups on the basis of DNA hybridization experiments, The polypeptides synthesized by the recombinants were predominantly LacZ fusion proteins, Serum obtained from patients who were clinically diagnosed to be in the early phase of M. tuberculosis infection was used to probe the 176 recombinants obtained. interestingly, some recombinants that gave very strong signals in the original screen did not react with early-phase serum; conversely, others whose signals were extremely weak in the original screen gave very intense signals with serum from recently infected patients, This indicates the differential nature of either the expression of these antigens or the immune response elicited by them as a function of disease progression.

The influence of HIV infection to the periodontium; a clinical, microbiological, and enzymological study

The main targets of human immunodeficiency virus (HIV) are CD4 receptors of CD4+ lymphocytes and many other cells such as monocytes/macrophages, megakaryocytes, peripheral blood dendritic cells, follicular dendritic cells (DC), epidermal Langerhans cells, and astrocytes. Infection and killing of CD4+ lymphocytes or false reaction of the body to HIV infection and the spontaneous apoptosis of CD4+ lymphocytes decrease CD4+ lymphocyte counts leading to immunosuppression, further disease progression, and appearance of opportunistic infections and malignancies. Oral manifestations are considered to be among the first signs of HIV infection. Enhanced degradation of extracellular matrix and basement membrane components in oral diseases including periodontitis is caused by Zn-dependent enzymes called matrix metalloproteinases (MMPs). The levels and degrees of activation of MMP-1, -2, -3, -7, -8, -9, -25, -26, tissue inhibitors of MMPs (TIMP)-1 and -2, and myeloperoxidase (MPO) and collagenolytic/gelatinolytic activities, and also Ig A, -G, and -M, total protein, and albumin levels in a two-year follow-up were studied from salivary samples. The expression of MMP-7, -8, -9, -25, and -26 immunoreactivities in gingival tissue specimens were studied. Healthy HIV-negative subjects served as controls. All studied clinical periodontal parameters and microbiological evaluation of the periodontopathogens showed that periodontal health of the HIV-positive patients was moderately decreased in comparison to the healthy controls. The levels of Candida in the periodontal pockets and salivary MPO increased with the severity of HIV infection. Immunoreactivities and levels of MMPs and TIMPs, and MMP activities (collagenase, gelatinase) were enhanced in the HIV-positive patient salivary samples relative to the healthy controls regardless of the phase of HIV infection. However, these parameters did not reflect periodontal status in a similar way as in the generally healthy periodontitis patients. Salivary total protein, albumin, IgA, -G, and -M levels were significantly higher in all phases of HIV infection compared to the controls, and salivary total protein, IgG and IgM levels remained higher after two years follow-up, partly correlating with the disease progression and which may reflect the leakage of serum components into the mouth and thus a decreased mucosal barrier. Salivary analyses of MMPs and TIMPs with immunohistochemical analyses showed that HIV infection could predispose to periodontal destruction when compared with healthy controls or the body s defence reactions associated with HIV infection may have been reflected or mediated by MMPs.

Periodontitis and peri-implantitis biomarkers in human oral fluids and the null-allele mouse model

Tissue destruction associated with the periodontal disease progression is caused by a cascade of host and microbial factors and proteolytic enzymes. Aberrant laminin-332 (Ln-332), human beta defensin (hBD), and matrix metalloproteinase (MMP) functions have been found in oral inflammatory diseases. The null-allele mouse model appears as the next step in oral disease research. The MMP-8 knock-out mouse model allowed us to clarify the involvement of MMP-8 in vivo in oral and related inflammatory diseases where MMP-8 is suggested to play a key role in tissue destruction. The cleaved Ln-332 γ2-chain species has been implicated in the apical migration of sulcular epithelial cells during the formation of periodontal pockets. We demonstrated that increased Ln-332 fragment levels in gingival crevicular fluid (GCF) are strongly associated with the severity of inflammation in periodontitis. Porphyromonas gingivalis trypsin-like proteinase can cleave an intact Ln-332 γ2-chain into smaller fragments and eventually promote the formation of periodontal pockets. hBDs are components of an innate mucosal defense against pathogenic microbes. Our results suggest that P. gingivalis trypsin-like proteinase can degrade hBD and thus reduce the innate immune response. Elevated levels and the increased activity of MMPs have been detected in several pathological tissue-destructive conditions where MMPs are shown to cleave extracellular matrix (ECM) and basement membrane (BM) molecules and to facilitate tissue destruction. Elevated levels of MMP-8 have been reported in many inflammatory diseases. In periodontitis, MMP-8 levels in gingival crevicular fluid (GCF) and in peri-implant sulcular fluid (PISF) are elevated at sites of active inflammation, and the increased levels of MMP-8 are mainly responsible for collagenase activity, which leads to tissue destruction. MMP-25, expressed by neutrophils, is involved in inflammatory diseases and in ECM turnover. MMP-26 can degrade ECM components and serve as an activator of other MMP enzymes. We further confirmed that increased levels and activation of MMP-8, -25, and -26 in GCF, PISF, and inflamed gingival tissue are associated with the severity of periodontal/peri-implant inflammation. We evaluated the role of MMP-8 in P. gingivalis-induced periodontitis by comparing MMP-8 knock-out (MMP8-/-) and wild-type mice. Surprisingly, MMP-8 significantly attenuated P. gingivalis-induced site-specific alveolar bone loss. We also evaluated systemic changes in serum immunoglobulin and lipoprotein profiles among these mouse groups. P. gingivalis infection increased HDL/VLDL particle size in the MMP-8-/- mice, which is an indicator of lipoprotein responses during systemic inflammation. Serum total LPS and IgG antibody levels were enhanced in both mice groups. P. gingivalis-induced periodontitis, especially in MMP-8-/- mice, is associated with severe alveolar bone loss and with systemic inflammatory and lipoprotein changes that are likely to be involved in early atherosclerosis.

Circulating tumor cells: Isolation, expansion, characterization and translation to clinic

Circulating tumor cells (CTCs) are the seeds for cancer metastases development, which is responsible for >90% of cancer-related deaths. Accurate quantification of CTCs in human fluids could be an invaluable tool for understanding cancer prognosis, delivering personalized medicine to prevent metastasis and finding cancer therapy effectiveness. Although CTCs were first discovered more than 200 years ago, until now it has been a nightmare for clinical practitioners to capture and diagnose CTCs in clinical settings. Our society needs rapid, sensitive, and reliable assays to identify the CTCs from blood in order to help save millions of lives. Due to the phenotypic EMT transition, CTCs are undetected for more than one-third of metastatic breast cancer patients in clinics. To tackle the above challenges, the first volume in “Circulating Tumor Cells (CTCs): Detection Methods, Health Impact and Emerging Clinical Challenges discusses recent developments of different technologies, which have the capability to target and elucidate the phenotype heterogenity of CTCS. It contains seven chapters written by world leaders in this area, covering basic science to possible device design which can have beneficial applications in society. This book is unique in its design and content, providing an in-depth analysis to elucidate biological mechanisms of cancer disease progression, CTC detection challenges, possible health effects and the latest research on evolving technologies which have the capability to tackle the above challenges. It describes the broad range of coverage on understanding CTCs biology from early predictors of the metastatic spread of cancer, new promising technology for CTC separation and detection in clinical environment and monitoring therapy efficacy via finding the heterogeneous nature of CTCs. (Imprint: Nova Biomedical)

Inflammation-driven bone formation in a mouse model of ankylosing spondylitis: Sequential not parallel processes

Background Ankylosing spondylitis (AS) is an immune-mediated arthritis particularly targeting the spine and pelvis and is characterised by inflammation, osteoproliferation and frequently ankylosis. Current treatments that predominately target inflammatory pathways have disappointing efficacy in slowing disease progression. Thus, a better understanding of the causal association and pathological progression from inflammation to bone formation, particularly whether inflammation directly initiates osteoproliferation, is required. Methods The proteoglycan-induced spondylitis (PGISp) mouse model of AS was used to histopathologically map the progressive axial disease events, assess molecular changes during disease progression and define disease progression using unbiased clustering of semi-quantitative histology. PGISp mice were followed over a 24-week time course. Spinal disease was assessed using a novel semi-quantitative histological scoring system that independently evaluated the breadth of pathological features associated with PGISp axial disease, including inflammation, joint destruction and excessive tissue formation (osteoproliferation). Matrix components were identified using immunohistochemistry. Results Disease initiated with inflammation at the periphery of the intervertebral disc (IVD) adjacent to the longitudinal ligament, reminiscent of enthesitis, and was associated with upregulated tumor necrosis factor and metalloproteinases. After a lag phase, established inflammation was temporospatially associated with destruction of IVDs, cartilage and bone. At later time points, advanced disease was characterised by substantially reduced inflammation, excessive tissue formation and ectopic chondrocyte expansion. These distinct features differentiated affected mice into early, intermediate and advanced disease stages. Excessive tissue formation was observed in vertebral joints only if the IVD was destroyed as a consequence of the early inflammation. Ectopic excessive tissue was predominantly chondroidal with chondrocyte-like cells embedded within collagen type II- and X-rich matrix. This corresponded with upregulation of mRNA for cartilage markers Col2a1, sox9 and Comp. Osteophytes, though infrequent, were more prevalent in later disease. Conclusions The inflammation-driven IVD destruction was shown to be a prerequisite for axial disease progression to osteoproliferation in the PGISp mouse. Osteoproliferation led to vertebral body deformity and fusion but was never seen concurrent with persistent inflammation, suggesting a sequential process. The findings support that early intervention with anti-inflammatory therapies will be needed to limit destructive processes and consequently prevent progression of AS.

Transforming growth factor -beta signaling through Smad3 in allergy : Studies in the mechanisms of asthma, atopic dermatitis and allergic contact reactions

Transforming growth factor β signalling through Smad3 in allergy Allergic diseases, such as atopic dermatitis, asthma, and contact dermatitis are complex diseases influenced by both genetic and environmental factors. It is still unclear why allergy and subsequent allergic disease occur in some individuals but not in others. Transforming growth factor (TGF)-β is an important immunomodulatory and fibrogenic factor that regulates cellular processes in injured and inflamed skin. TGF-β has a significant role in the regulation of the allergen-induced immune response participating in the development of allergic and asthmatic inflammation. TGF-β is known to be an immunomodulatory factor in the progression of delayed type hypersensitivity reactions and allergic contact dermatitis. TGF-β is crucial in regulating the cellular responses involved in allergy, such as differentiation, proliferation and migration. TGF-β signals are delivered from the cytoplasm to the nucleus by TGF-β signal transducers called Smads. Smad3 is a major signal transducer in TGF-β -signalling that controls the expression of target genes in the nucleus in a cell-type specific manner. The role of TGF-β-Smad3 -signalling in the immunoregulation and pathophysiology of allergic disorders is still poorly understood. In this thesis, the role of TGF-β-Smad -signalling pathway using Smad3 -deficient knock out mice in the murine models of allergic diseases; atopic dermatitis, asthma and allergic contact reactions, was examined. Smad3-pathway regulates allergen induced skin inflammation and systemic IgE antibody production in a murine model atopic dermatitis. The defect in Smad3 -signalling decreased Th2 cytokine (IL-13 and IL-5) mRNA expression in the lung, modulated allergen induced specific IgG1 response, and affected mucus production in the lung in a murine model of asthma. TGF-β / Smad3 -signalling contributed to inflammatory hypersensitivity reactions and disease progression via modulation of chemokine and cytokine expression and inflammatory cell recruitment, cell proliferation and regulation of the specific antibody response in a murine model of contact hypersensitivity. TGF-β modulates inflammatory responses - at least partly through the Smad3 pathway - but also through other compensatory, non-Smad-dependent pathways. Understanding the effects of the TGF-β signalling pathway in the immune system and in disease models can help in elucidating the multilevel effects of TGF-β. Unravelling the mechanisms of Smad3 may open new possibilities for treating and preventing allergic responses, which may lead to severe illness and loss of work ability. In the future the Smad3 signalling pathway might be a potential target in the therapy of allergic diseases.

Juvenile neuronal ceroid lipofuscinosis; psychiatric symptoms, GAD-autoantibodies, and response to medication

Juvenile neuronal ceroid lipofuscinosis (JNCL) is one of the most common neurodegenerative diseases in childhood. Its clinical onset, with visual failure as the first sign, is between the ages of 4 to 8 years. During the disease progress, epilepsy, motor symptoms, cognitive decline, and psychiatric symptoms become apparent. It leads to premature death between ages 15 and 30. Treatment consists of symptomatic drug administration and various forms of rehabilitation, but to date, no curative treatment exists. To gain a more comprehensive picture of psychiatric problems, symptoms were evaluated by the Child Behavior Checklist, the Teacher Report Form, and the Children s Depression Inventory. The JNCL patients had a great number of severe psychiatric symptoms, with wide inter-individual variability. The most common symptoms were social, thought, attention, and sleep problems, somatic complaints, and aggressive behaviour. Patients with psychotropic treatment had more problems than did those without psychotropic treatment, and female patients had more problems than did males. Between 10 and 20% of the patients reported depressive symptoms. In a 5-year follow-up, [123I]β-CIT SPECT and MRI revealed a tendency of decreasing serotonin transporter (SERT) availability and progressive brain atrophy. The correlation between changes in midbrain SERT and total brain volume was positive; no correlation appeared between SERT or brain atrophy and depressive symptoms. Thus, it seems likely that the low SERT availability is associated with progressive brain atrophy; it may also predispose towards depression, however. An open survey of psychotropic drugs and their efficacy was performed on JNCL patients in Finland. The most commonly used psychotropic drugs were the antidepressant citalopram and the antipsychotic risperidone. Their efficacy was good or satisfactory in the majority of cases and they seemed well tolerated. Quetiapine had a marked effect on one patient with a history of severe psychotic symptoms. Glutamate decarboxylase 65 autoantibodies (GAD65ab), found in JNCL patients, indicate that an immunomediated reaction against GAD or GABAergic neurons may play a part in the underlying pathogenetic mechanism. GAD65ab s also appeared in the serum of all eight JNCL patients included and intermittent corticosteroid therapy was initiated in all cases. After one year, the GAD65ab s had disappeared in the two oldest patients, who experienced an improvement in motor symptoms and alertness associated with their prednisolone therapy. Two younger patients experienced a significant IQ increase, but no change in GADab s. A randomized study with longer follow-up time is needed, however, to clarify the effect of prednisolone on disease progression.

Characterisation of novel primary tumour derived epithelial prostate cancer cell lines

Current translational and basic prostate cancer research is limited by the number of cell lines that truly reflect the spectrum of disease progression, with most commonly used cell lines being derived from metastatic lesions. There are essentially no prostate cancer cell lines derived from primary tumours or localised disease in wide use.

HIV outbreak among injecting drug users in Finland

An HIV outbreak among Finnish injecting drug users (IDUs) occurred in 1998. By the end of 2005, 282 IDUs were in-fected, most of them by recombinant virus CRF01_AE of HIV. After a rapid spread, the outbreak subsided, and the prevalence of HIV among IDUs remained low (<2%). The purpose of the study was to describe the outbreak in order to recognise factors that have influenced the spread and restriction of the outbreak, and thus to find tools for HIV preven-tion. Data on Finnish IDUs newly diagnosed HIV-positive between 1998 and 2005 was collected through interviews and patient documents. Study I compared markers of disease progression between 93 Finnish IDUs and 63 Dutch IDUs. In study II, geographical spread of the HIV outbreak was examined and compared with the spatial distribution of employed males. In study III, risk behaviour data from interviews of 89 HIV-positive and 207 HIV-negative IDUs was linked, and prevalence and risk factors for unprotected sex were evaluated. In study IV, data on 238 newly diagnosed IDUs was combined with data on 675 sexually transmitted HIV cases, and risk factors for late HIV diagnosis (CD4 cell count <200/µL, or AIDS at HIV diagnosis) were analysed. Finnish IDUs infected with CRF01_AE exhibited higher viral loads than did Amsterdam IDUs infected with subtype B, but there was no difference in CD4 development. The Finnish IDU outbreak spread and was restricted socially in a marginalised IDU population and geographically in areas characterised by low proportions of employed males. Up to 40% of the cases in the two clusters outside the city centre had no contact with the centre, where needle exchange services were available since 1997. Up to 63% of HIV-positive and 80% of HIV-negative sexually active IDUs reported inconsistent condom use, which was associated with steady relationships and recent inpatient addiction care. Com-pared to other transmission groups, HIV-positive IDUs were diagnosed earlier in their infection. The proportion of late diagnosed HIV cases in all transmission groups was 23%, but was only 6% among IDUs diagnosed during the first four years of the epidemic. The high viral load in early HIV infection may have contributed to the rapid spread of recombinant virus in the Finnish outbreak. The outbreak was restricted to a marginalised IDU population, and limited spatially to local pockets of pov-erty. To prevent HIV among IDUs, these pockets should be recognised and reached early through outreach work and the distribution of needle exchange and other prevention activities. To prevent the sexual transmission of HIV among IDUs, prevention programmes should be combined with addiction care services and targeted at every IDU. The early detection of the outbreak and early implementation of needle exchange programmes likely played a crucial role in re-versing the IDU outbreak.

Afatinib versus gefitinib as first-line treatment of patients with EGFR mutation-positive non-small-cell lung cancer (LUX-Lung 7): a phase 2B, open-label, randomised controlled trial

Background The irreversible ErbB family blocker afatinib and the reversible EGFR tyrosine kinase inhibitor gefitinib are approved for first-line treatment of EGFR mutation-positive non-small-cell lung cancer (NSCLC). We aimed to compare the efficacy and safety of afatinib and gefitinib in this setting. Methods This multicentre, international, open-label, exploratory, randomised controlled phase 2B trial (LUX-Lung 7) was done at 64 centres in 13 countries. Treatment-naive patients with stage IIIB or IV NSCLC and a common EGFR mutation (exon 19 deletion or Leu858Arg) were randomly assigned (1:1) to receive afatinib (40 mg per day) or gefitinib (250 mg per day) until disease progression, or beyond if deemed beneficial by the investigator. Randomisation, stratified by EGFR mutation type and status of brain metastases, was done centrally using a validated number generating system implemented via an interactive voice or web-based response system with a block size of four. Clinicians and patients were not masked to treatment allocation; independent review of tumour response was done in a blinded manner. Coprimary endpoints were progression-free survival by independent central review, time-to-treatment failure, and overall survival. Efficacy analyses were done in the intention-to-treat population and safety analyses were done in patients who received at least one dose of study drug. This ongoing study is registered with ClinicalTrials.gov, number NCT01466660. Findings Between Dec 13, 2011, and Aug 8, 2013, 319 patients were randomly assigned (160 to afatinib and 159 to gefitinib). Median follow-up was 27·3 months (IQR 15·3–33·9). Progression-free survival (median 11·0 months [95% CI 10·6–12·9] with afatinib vs 10·9 months [9·1–11·5] with gefitinib; hazard ratio [HR] 0·73 [95% CI 0·57–0·95], p=0·017) and time-to-treatment failure (median 13·7 months [95% CI 11·9–15·0] with afatinib vs 11·5 months [10·1–13·1] with gefitinib; HR 0·73 [95% CI 0·58–0·92], p=0·0073) were significantly longer with afatinib than with gefitinib. Overall survival data are not mature. The most common treatment-related grade 3 or 4 adverse events were diarrhoea (20 [13%] of 160 patients given afatinib vs two [1%] of 159 given gefitinib) and rash or acne (15 [9%] patients given afatinib vs five [3%] of those given gefitinib) and liver enzyme elevations (no patients given afatinib vs 14 [9%] of those given gefitinib). Serious treatment-related adverse events occurred in 17 (11%) patients in the afatinib group and seven (4%) in the gefitinib group. Ten (6%) patients in each group discontinued treatment due to drug-related adverse events. 15 (9%) fatal adverse events occurred in the afatinib group and ten (6%) in the gefitinib group. All but one of these deaths were considered unrelated to treatment; one patient in the gefitinib group died from drug-related hepatic and renal failure. Interpretation Afatinib significantly improved outcomes in treatment-naive patients with EGFR-mutated NSCLC compared with gefitinib, with a manageable tolerability profile. These data are potentially important for clinical decision making in this patient population.