Mesothelin antigen density influences anti-mesothelin chimeric antigen receptor T cell cytotoxicity.

BACKGROUND AIMS: Several anti-mesothelin (MSLN) chimeric antigen receptor (CAR) T cells are in phase 1/2 clinical trials to treat solid-organ malignancies. The effect of MSLN antigen density on MSLN CAR cytotoxicity against tumor cells has not been examined previously, nor are there data regarding the effect of agents that increase MSLN antigen density on anti-MSLN CAR T cell efficacy. METHODS: MSLN antigen density was measured on a panel of pancreatic cancer and mesothelioma cell lines by flow cytometry. In parallel, the cytotoxicity and specificity of two anti-MSLN CAR T cells (m912 and SS1) were compared against these cell lines using a real-time impedance-based assay. The effect of two MSLN 'sheddase' inhibitors (lanabecestat and TMI-1) that increase MSLN surface expression was also tested in combination with CAR T cells. RESULTS: SS1 CAR T cells were more cytotoxic compared with m912 CAR T cells against cell lines that expressed fewer than ∼170 000 MSLN molecules/cell. A comparison of the m912 and amatuximab (humanized SS1) antibodies identified that amatuximab could detect and bind to lower levels of MSLN on pancreatic cancer and mesothelioma cell lines, suggesting that superior antibody/scFv affinity was the reason for the SS1 CAR's superior cytotoxicity. The cytotoxicity of m912 CAR T cells was improved in the presence of sheddase inhibitors, which increased MSLN antigen density. CONCLUSIONS: These data highlight the value of assessing CAR constructs against a panel of cells expressing varying degrees of target tumor antigen as occurs in human tumors. Furthermore, the problem of low antigen density may be overcome by concomitant administration of drugs that inhibit enzymatic shedding of MSLN.

The 4-1BBζ costimulatory domain in chimeric antigen receptors enhances CD8+ T-cell functionality following T-cell receptor stimulation.

BACKGROUND: Chimeric antigen receptor (CAR) T-cells have revolutionized the treatment of CD19- and B-cell maturation antigen-positive haematological malignancies. However, the effect of a CAR construct on the function of T-cells stimulated via their endogenous T-cell receptors (TCRs) has yet to be comprehensively investigated. METHODS: Experiments were performed to systematically assess TCR signalling and function in CAR T-cells using anti-mesothelin human CAR T-cells as a model system. CAR T-cells expressing the CD28 or 4-1BB costimulatory endodomains were manufactured and compared to both untransduced T-cells and CAR T-cells with a non-functional endodomain. These cell products were treated with staphylococcal enterotoxin B to stimulate the TCR, and in vitro functional assays were performed by flow cytometry. RESULTS: Increased proliferation, CD69 expression and IFNγ production were identified in CD8+ 4-1BBζ CAR T-cells compared to control untransduced CD8+ T-cells. These functional differences were associated with higher levels of phosphorylated ZAP70 after stimulation. In addition, these functional differences were associated with a differing immunophenotype, with a more than two-fold increase in central memory cells in CD8+ 4-1BBζ CAR T-cell products. CONCLUSION: Our data indicate that the 4-1BBζ CAR enhances CD8+ TCR-mediated function. This could be beneficial if the TCR targets epitopes on malignant tissues or infectious agents, but detrimental if the TCR targets autoantigens.

Use of adrenaline to manage suspected anaphylaxis following COVID-19 vaccination: An Australian retrospective cohort study.

BACKGROUND: The rate of anaphylaxis following COVID-19 vaccinations is estimated to be 2-11 cases per million doses administered. However, adrenaline is occasionally used in individuals who are later diagnosed with immunisation stress-related responses, as their initial presenting signs and symptoms can appear similar to that of anaphylaxis. This study aims to describe the clinical profile of individuals who had received adrenaline following a COVID-19 vaccine and their subsequent revaccination outcomes. METHODS: We examined notifications of cases who had received adrenaline following a COVID-19 vaccine in New South Wales, Australia. The cases were classified into Brighton Collaboration Case Definition (BCCD) for anaphylaxis, their clinical presentation, management and subsequent revaccination outcomes were compared. RESULTS: From 22 February 2021 to 30 September 2021, there were 222 cases where adrenaline was administered. Of these, 32 (14 %) fulfilled Level 1 BCCD, 59 (27%) Level 2, 2 (1%) Level 3, 97 (44%) Level 4 and 32 (14 %) Level 5. The most commonly reported symptoms were sensation of throat closure (n = 116, 52%), difficulty breathing (n = 82, 37%) and nausea (n = 55, 25 %). Of the 176 (79%) individuals who proceeded to further vaccination, 89 (51%) received the same vaccine formulation and only 14 (8%) experienced another allergic adverse event with 9 (5%) receiving adrenaline. CONCLUSION: Less than one in five individuals who received adrenaline met Level 1 BCCD criteria for anaphylaxis. Many reactions that were treated with adrenaline had little to no diagnostic certainty of anaphylaxis and in such cases repeat vaccination had a high likelihood of being tolerated. Increased awareness and education on objective signs and symptoms of anaphylaxis is required to ensure appropriate use of adrenaline.

High mesothelin expression by immunohistochemistry predicts improved survival in pleural mesothelioma.

AIMS: Mesothelin (MSLN) is a cancer-associated antigen that is overexpressed in malignancies such as mesothelioma, pancreatic and ovarian cancer. It is also a target for novel personalised therapies, including antibodies, antibody-drug conjugates and chimeric antigen receptor T cells. Immunohistochemistry may predict those who would best respond to anti-mesothelin therapies and guide decisions in therapeutic strategy. This study aimed to assess the intensity and distribution of MSLN immunostaining in mesothelioma, and to determine the prognostic value of MSLN expression by histochemical-score (H-score). METHODS AND RESULTS: The MN1 anti-MSLN antibody was used to stain a formalin-fixed paraffin-embedded tissue microarray of histologically confirmed mesothelioma from 75 consecutive patients who had undergone pleurectomy with or without decortication. MSLN positivity, the staining intensity, distribution of staining and H-score were evaluated. The correlation of H-score with prognosis was investigated. Sixty-six per cent of epithelioid tumours were MSLN-positive (with expression in > 5% tumour cells). Of MSLN-expressing epithelioid tumours, 70.4% had moderate (2+) or strong (3+) intensity MSLN immunostaining, although only 37% of samples had staining in ≥ 50% of tumour cells. In multivariate analysis, MSLN H-score as a continuous variable and an H-score ≥ 33 were independent predictors of improved survival (P = 0.04 and P < 0.001, respectively). CONCLUSIONS: MSLN expression was more heterogenous in epithelioid mesothelioma than reported previously. Therefore, it would be appropriate to perform an immunohistochemical assessment of MSLN expression to stratify and assess patient suitability for mesothelin-targeted personalised therapies, such as chimeric antigen receptor T cells.

A Composite Serum Biomarker Index for the Diagnosis of Systemic Sclerosis-Associated Interstitial Lung Disease: A Multicenter, Observational Cohort Study.

OBJECTIVE: In patients with systemic sclerosis (SSc), we investigated composite serum biomarker panels for the diagnosis and risk stratification of SSc-associated interstitial lung disease (SSc-ILD). METHODS: We analyzed 28 biomarkers in 640 participants: 259 patients with SSc-ILD and 179 SSc patients without ILD (Australian Scleroderma Cohort Study), 172 patients with idiopathic pulmonary fibrosis (IPF-controls) (Australian IPF Registry), and 30 healthy controls. A composite index was developed from biomarkers associated with ILD in multivariable analysis derived at empirical thresholds. We evaluated the performance of the index to identify ILD, and specifically SSc-ILD, and its association with lung function, disease extent on radiography, and patient health-related quality of life in derivation and validation cohorts. Biomarkers to distinguish SSc-ILD from IPF-controls were identified. RESULTS: A composite biomarker index, comprising surfactant protein D (SP-D), Ca15-3, and intercellular adhesion molecule 1 (ICAM-1), was strongly associated with SSc-ILD diagnosis, independent of age, sex, smoking history, and lung function (for biomarker index score 3, pooled adjusted odds ratio was 12.72 (95% confidence interval 4.59-35.21) (P < 0.001). The composite index strengthened the performance of individual biomarkers for SSc-ILD identification. In SSc patients, a higher index was associated with worse baseline disease severity (for biomarker index score 3 relative to biomarker index score 0, the adjusted absolute change in forced vital capacity percent predicted was -17.84% and the diffusing capacity for carbon monoxide percent predicted was -20.16%; both P < 0.001). CONCLUSION: A composite serum biomarker index, comprising SP-D, Ca15-3, and ICAM-1, may improve the identification and risk stratification of ILD in SSc patients at baseline.

Hemophagocytic lymphohistiocytosis (HLH) and cytokine release syndrome (CRS) in a patient with oncogene-addicted metastatic non-small cell lung cancer (NSCLC) following combination chemotherapy-immunotherapy.

Immune checkpoint inhibitors (ICIs) are utilized in a variety of clinical settings for the management of patients with metastatic non-small cell lung cancer (mNSCLC). While any organ may be subject to immune-related adverse events (irAEs) as a consequence of ICI therapy, hematological irAEs are uncommon. We describe a scenario involving a patient with oncogene-addicted mNSCLC who experienced the rare, life-threatening complication of hemophagocytic lymphohistiocytosis (HLH) and cytokine release syndrome following the receipt of the IMPower150 regimen (carboplatin/paclitaxel/atezolizumab/bevacizumab) after progression on initial tyrosine kinase inhibitor therapy. Malignancy-associated HLH, while previously described, is more typically associated with hematological rather than solid cancers and has only very recently been reported among patients receiving ICIs. While identification of hemophagocytosis on bone marrow examination is pathognomonic, this feature is not essential for confirming a diagnosis of HLH. Prompt recognition of suspicious laboratory and clinical features by medical oncologists and engagement with other relevant disciplines is hence critical to ensure optimal management of the condition.

Gluten-free diet adherence and implications for the diagnosis of coeliac disease.

Coeliac disease (CD) is an autoimmune disorder caused by the ingestion of gluten containing foods in genetically susceptible individuals, with a worldwide prevalence of up to 1%. Currently, the only available treatment is a gluten-free diet (GFD). Screening for CD is primarily performed using serum based testing for anti-tissue transglutaminase (tTG) antibodies. Patients must be on a gluten containing diet at the time of testing to ensure an accurate serological result. We investigated the prevalence of a GFD in hospital clinic settings and the general population using survey data to estimate the proportion of CD patients that may be misdiagnosed for CD based on serological tests. Data were collected at clinics of a metropolitan hospital in Sydney, Australia, and the general population. Data from Medicare Benefits Scheme and tTG results from a large Australian private laboratory were reviewed for comparison. Of 778 participants who responded to the survey, 58 (7.5%) were on a GFD. More patients attending the immunology (15.9%) and gastroenterology (12.1%) clinics adopted a GFD than those attending the diabetes (2.6%) or endocrinology (6.1%) clinics, or in the general population (4.3%). More females than males excluded gluten from their diet (p<0.0001). Medicare statistics between 2013 and 2019 demonstrated an increase in CD serological testing; however, tTG data from a private pathology highlighted a stable level of elevated tTG antibodies of 3% of total tests performed. The high number of individuals on a GFD is likely impacting the ability to accurately diagnose CD using serum-based testing.

Subunit-specific autoantibodies in autoimmune autonomic ganglionopathy.

Autoimmune Autonomic Ganglionopathy (AAG) is a disorder that causes autonomic failure and is associated with alpha3-ganglionic acetylcholine-receptor (gnACHR) antibodies. Assays that detect antibodies to whole gnACHR or subunits are available. We compared in-house subunit-specific immunoassays using bacterially-expressed alpha3 and beta4 subunits against an immunomodulation assay to detect antibodies in patients with AAG or control groups in a novel 2-step clinical-characteristic unblinding protocol. Only 1/8 patients with seropositive-AAG had subunit-specific antibodies, with sensitivity, specificity, false-negative and positive rates of 12.5, 85.2, 78.6 and 13.4% respectively. Subunit-specific antibody-derived false-positive results can lead to misdiagnosis, as autonomic failure is not specific to AAG.

Autoimmune autonomic ganglionopathy: Ganglionic acetylcholine receptor autoantibodies.

Autoimmune Autonomic Ganglionopathy (AAG) is a rare immune-mediated disease of the autonomic nervous system. The incidence of AAG is unknown and diagnosis is often difficult due to the multicompartmental nature of the autonomic nervous system - sympathetic, parasympathetic and enteric components - with variable severity and number of components affected. Diagnostic confidence is increased when ganglionic acetylcholine receptor (gnACHR) autoantibodies are detected. Three gnACHR autoantibody diagnostic assays have been described (two binding assays, one receptor immunomodulation assay), but cross-validation between assays is limited. The prevalence of gnACHR autoantibodies in AAG is not known, with application of different clinical and laboratory criteria in the few studies of AAG cohorts and large retrospective laboratory studies of positive gnACHR autoantibodies lacking adequate clinical characterisation. Furthermore, the rate of unexpected gnACHR autoantibody positivity in conditions without overt autonomic dysfunction (false positive results) adds to the complexity of their interpretation. We review the pathophysiology of gnACHR autoantibodies and assays for their detection, with immunomodulation and high titer radioimmunoprecipitation results likely offering better AAG disease identification.

Evaluation of commercially available antibodies and fluorescent conotoxins for the detection of surface ganglionic acetylcholine receptor on the neuroblastoma cell line, IMR-32 by flow cytometry.

Commercially available antibodies that bind to the human muscle acetylcholine receptor (ACHR) have been validated previously for flow cytometric use (Keefe et al., 2009; Leite et al., 2008; Lozier et al., 2015). Despite a multitude of commercially available antibodies to other nicotinic ACHRs, validation in a wide variety of immunoassay formats is lacking; when studied, a large proportion of these antibodies have been deemed not fit for most research purposes (Garg and Loring, 2017). We have recently described a flow cytometric immunomodulation assay for the diagnosis of Autoimmune Autonomic Ganglionopathy (AAG) (Urriola et al., 2021) that utilises the monoclonal antibody mab35(Urriola et al., 2021) which is specific for ganglionic ACHR (gnACHR) that contain α3 subunits (Vernino et al., 1998). Other fluorescent ligands for α3-gnACHR have not been validated for flow cytometric use. We investigated 7 commercially sourced antibodies and 3 synthetic fluorescent novel conotoxins purported to specifically bind to the extracellular domains of the gnACHR, and compared the results to staining by mab35, using flow cytometry with the neuroblastoma cell line IMR-32. We also evaluated the degree of non-specific binding by depleting the cell membrane of the relevant acetylcholine receptor with a pre-incubation step involving the serum from a patient with Autoimmune Autonomic Ganglionopathy containing pathogenic antibodies to the ganglionic acetylcholine receptor. None of the assessed conotoxins, and only one antibody (mab35) was found to perform adequately in flow cytometric staining of the native ganglionic acetylcholine receptor.

Diagnosis of myositis-associated interstitial lung disease: Utility of the myositis autoantibody line immunoassay.

OBJECTIVES: The detection of myositis autoantibodies (MA) in patients with interstitial lung disease (ILD) has major implications for diagnosis and management, especially amyopathic and forme frustes of idiopathic inflammatory myositis-associated ILD (IIM-ILD). Use of the MA line immunoblot assay (MA-LIA) in non-rheumatological cohorts remains unvalidated. We assessed the diagnostic performance of the MA-LIA and explored combined models with clinical variables to improve identification of patients with IIM-ILD. METHODS: Consecutive patients referred to a specialist ILD clinic, with ILD-diagnosis confirmed at multidisciplinary meeting, and MA-LIA performed within six months of baseline were included. Pre-specified MA-LIA thresholds were evaluated for IIM-ILD diagnosis. RESULTS: A total 247 ILD patients were included (IIM-ILD n = 12, non-IIM connective tissue disease-associated ILD [CTD-ILD] n = 52, idiopathic interstitial pneumonia [IIP] n = 115, other-ILD n = 68). Mean age was 64.8 years, with 45.3% female, mean FVC 75.5% and DLCO 59.2% predicted. MA were present in 13.8% overall and 83.3% of IIM-ILD patients. The most common MA in IIM-ILD and non-IIM ILD patients were anti-Jo-1 (prevalence 40%) and anti-PMScl (29.2%) autoantibodies respectively. The pre-specified low-positive threshold (>10 signal intensity) had the highest discriminative capacity for IIM-ILD (AUC 0.86). Combining MA-LIA with age, gender, clinical CTD-manifestations and an overlap non-specific interstitial pneumonia/organising pneumonia pattern on HRCT improved discrimination for IIM-ILD (AUC 0.96). CONCLUSION: The MA-LIA is useful to support a diagnosis of IIM-ILD as a complement to multi-disciplinary ILD assessment. Clinical interpretation is optimised by consideration of the strength of the MA-LIA result together with clinical and radiological features of IIM-ILD.

A Flow Cytometric Assay to Detect Functional Ganglionic Acetylcholine Receptor Antibodies by Immunomodulation in Autoimmune Autonomic Ganglionopathy.

Autoimmune Autonomic Ganglionopathy (AAG) is an uncommon immune-mediated neurological disease that results in failure of autonomic function and is associated with autoantibodies directed against the ganglionic acetylcholine receptor (gnACHR). The antibodies are routinely detected by immunoprecipitation assays, such as radioimmunoassays (RIA), although these assays do not detect all patients with AAG and may yield false positive results. Autoantibodies against the gnACHR exert pathology by receptor modulation. Flow cytometric analysis is able to determine if this has occurred, in contrast to the assays in current use that rely on immunoprecipitation. Here, we describe the first high-throughput, non-radioactive flow cytometric assay to determine autoantibody mediated gnACHR immunomodulation. Previously identified gnACHR antibody seronegative and seropositive sera samples (RIA confirmed) were blinded and obtained from the Oxford Neuroimmunology group along with samples collected locally from patients with or without AAG. All samples were assessed for the ability to cause gnACHR immunomodulation utilizing the prototypical gnACHR expressing cell line, IMR-32. Decision limits were calculated from healthy controls, and Receiver Operating Characteristic (ROC) curves were constructed after unblinding all samples. One hundred and ninety serum samples were analyzed; all 182 expected negative samples (from healthy controls, autonomic disorders not thought to be AAG, other neurological disorders without autonomic dysfunction and patients with Systemic Lupus Erythematosus) were negative for immunomodulation (<18%), as were the RIA negative AAG and unconfirmed AAG samples. All RIA positive samples displayed significant immunomodulation. There were no false positive or negative samples. There was perfect qualitative concordance as compared to RIA, with an Area Under ROC of 1. Detection of Immunomodulation by flow cytometry for the identification of gnACHR autoantibodies offers excellent concordance with the gnACHR antibody RIA, and overcomes many of the shortcomings of immunoprecipitation assays by directly measuring the pathological effects of these autoantibodies at the cellular level. Further work is needed to determine the correlation between the degree of immunomodulation and disease severity.

Baseline Characteristics and Survival of an Australian Interstitial Pneumonia with Autoimmune Features Cohort.

BACKGROUND AND OBJECTIVE: The research term "interstitial pneumonia with autoimmune features" (IPAF) encompasses interstitial lung disease (ILD) patients with autoimmune features not meeting diagnostic criteria for a defined connective tissue disease (CTD). It remains unclear if IPAF is a distinct disease entity with implications for management and prognosis. We describe an Australian IPAF population and compare their baseline characteristics and outcomes with distinct cohorts of idiopathic interstitial pneumonia (IIP), CTD-ILD, and unclassifiable ILD. METHODS: Review of 291 consecutive patients attending a specialist ILD clinic was performed. Patients with a diagnosis of IIP, CTD-ILD, and unclassifiable ILD by ILD-multidisciplinary meeting (ILD-MDM) were included. Patients meeting the IPAF criteria were identified. Baseline clinical data, survival, and progression were compared between ILD groups. RESULTS: 226 patients were included, 36 meeting the IPAF criteria. IPAF patients demonstrated a high prevalence of autoantibodies to tRNA synthetase (35.3%), Ro52 (27.8%), and neutrophilic cytoplasmic antigens (ANCA; 20.0%). IPAF and CTD-ILD patients demonstrated similar clinical characteristics (mean age 66.6 and 63.7 years, respectively, female predominant, frequent CTD-manifestations). Lung function did not differ between ILD groups. Disease severity, pulmonary hypertension (PH), and ILD-MDM diagnosis were strong predictors of worse transplant-free survival (TFS). Meeting the IPAF criteria was not associated with TFS. CONCLUSIONS: We identified IPAF as a heterogeneous phenotype that overlaps considerably with CTD-ILD. Disease severity, PH, and ILD-MDM diagnosis were more powerful predictors of survival outcomes than meeting the IPAF criteria.

Cerebrospinal fluid free light chain quantitation is a specific biomarker for inflammatory neurological disorders in a paediatric patient cohort.

The analysis of cerebrospinal fluid (CSF) is routinely used in the diagnostic work-up of a range of inflammatory, infective, and congenital neurological conditions. Many diagnostic tests used in this analysis have poor sensitivity; as such, we investigated the utility of CSF free light chain (FLC) analysis as an adjunct to currently used assays in a paediatric population with neurological disorders. Kappa (κ) and lambda (λ) FLC levels were quantitated in blinded CSF samples by two nephelometric platforms. Results were correlated to clinical diagnoses and classified according to inflammatory/infective or non-inflammatory pathogenesis. FLC results were also compared to currently used CSF diagnostic tests including oligoclonal bands (OCB), CSF IgG and albumin levels, and differential cell count. Of 70 samples analysed, 29 (41%) had an inflammatory or infective diagnosis and 41 (59%) presented with a range of non-inflammatory aetiologies. Thirteen patients had elevated κFLC or λFLC as detected on the IMMAGE 800, defined as greater than the detection limit of the assay (0.600 mg/L for CSF κFLC, and 0.490 mg/L for CSF λFLC), and of these 12 (92%) had an inflammatory disease (sensitivity 41.4%, specificity 97.6%). On the BN II using optimal cut-offs of 0.27 mg/L and 0.12 mg/L for CSF κFLC and λFLC respectively, 24 (34%) patients had elevated results, of which 21 (88%) had an inflammatory disease (sensitivity 72.4%, specificity 92.7%). Analysis of FLC correlated better with diagnostic classification of the diseases than OCB, cell counts and CSF IgG levels. The results of this study support the use of CSF FLC analysis in the diagnosis of paediatric neuroinflammatory conditions.

Precision therapy for neuromyelitis optica spectrum disorder: A retrospective analysis of the use of class-switched memory B-cells for individualised rituximab dosing schedules.

BACKGROUND: B-cell depleting treatments are widely used to modify the course of neuromyelitis optica spectrum disorder (NMOSD). Despite recent successful Phase 3 trials of several novel NMOSD therapies, limited availability and high cost constrains their clinical use, and rituximab (RTX) remains a core treatment in many centres. Since 2013, the Royal Prince Alfred Hospital Neuroimmunology Clinic (NIC) has regularly measured class-switched memory B-cells (SMB-cells) in the peripheral blood of patients with NMOSD, who have been treated with RTX, in order to guide retreatment intervals. OBJECTIVE: To assess the management and outcomes of the treated patients, and to determine the effect of SMB-cell monitoring in guiding retreatment intervals. METHODS: A retrospective analysis of hospital records, clinic letters and laboratory data was performed. RESULTS: Sixteen patients with NMOSD received individualised rituximab dosing at NIC between 2013 and 2018. Fourteen (87.5%) were aquaporin-4 antibody (AQP4-Ab) positive; 1 (6.25%) was myelin oligodendrocyte glycoprotein antibody (MOG-Ab) positive and 1 (6.25%) was seronegative. After commencement of RTX, individually dosed according to regular measurements of serum SMB-cells, there was a 77.5% reduction in annualised relapse rate over a mean follow-up time of 46.1 months in our recently active NMOSD patients. Their mean retreatment interval was 50.9 weeks. CONCLUSIONS: This study provides real-world evidence supporting individualised rituximab dosing in the treatment of NMOSD.

Liver enzyme profile and progression in association with thyroid autoimmunity in Graves' disease.

OBJECTIVE: To investigate the associations of Graves' disease (GD) severity, autoimmunity and longitudinal liver enzyme changes with time in a cohort with well-characterized GD. DESIGN: Retrospective cohort study. PATIENTS: Patients diagnosed with Graves' disease, treated at Royal Prince Alfred Hospital Sydney, Adult Thyroid Clinic from 2000 to 2012 inclusive. MEASUREMENTS: Inclusion criteria were patients with a complete set of TSH, FT4, FT3, liver enzymes and TSH receptor antibody (TRAb) results prior to commencement of thionamide therapy. RESULTS: Of the 146 patients who had complete results, 69 (47%) had at least one abnormal liver enzyme. Gamma glutamyltransferase (GGT) was most frequently abnormal (74%), followed by alanine aminotransferase (ALT) (57%), alkaline phosphatase (ALP) (39%) and then aspartate aminotransferase (AST) (29%). Subsequent to thyroid function normalization, 78% of the liver enzymes were normalized, 10% were persistently abnormal and 12% were lost to follow-up. Circulating TRAb, FT3 and FT4 results were categorized into mild, moderate and severe elevations. At univariate regression analyses, TRAb, FT3 and FT4 levels were each significantly associated with abnormal liver enzyme profile. Multivariate regression including TRAB, FT3 and FT4 as independent variables demonstrated FT3 and FT4 were more strongly associated with abnormal liver profile than TRAb. However, the initial FT3 and FT4 levels were not associated with abnormal liver profile in the subgroup with persistently abnormal liver profile. CONCLUSION: Graves' disease is commonly associated with abnormal liver enzymes, and most commonly with abnormal levels of GGT, and that an abnormal liver enzyme profile is more directly linked to the degree of thyrotoxicosis than levels of TRAB.

Review: Serum biomarkers in idiopathic pulmonary fibrosis and systemic sclerosis associated interstitial lung disease - frontiers and horizons.

Disease behaviour in interstitial lung disease (ILD) is highly variable and accurate clinical tools to predict prognosis and guide management decisions remain unsatisfactorily elusive. Accurate disease stratification would allow clinicians to better distinguish patients at risk of rapid progression requiring urgent treatment, from those indolent disease where potentially toxic drug therapy could be minimised or avoided. Several serum biomarkers have demonstrated potential utility for diagnosis and prognosis of ILD in small retrospective studies, and the hope is future multicentre prospective trials focussed on the markers with most potential will see translation to clinical practice. Two important and contrasting fibrotic lung diseases with high mortality are idiopathic pulmonary fibrosis (IPF) and systemic sclerosis associated ILD (SSc-ILD). In this era where anti-fibrotics for IPF have proven benefit, there are increasing biologic and non-biologic options for the treatment of connective tissue disease ILD (CTD-ILD), and the incidence of both is increasing, there is an urgent need to improve the diagnostic and prognostic accuracy in these complex patients. This comprehensive literature review will summarise and discuss the current evidence for the major candidate serum biomarkers in IPF and SSc-ILD. Biomarkers will be categorised by the following major mechanistic pathways (1) alveolar epithelial cell damage; (2) aberrant fibrogenesis, fibroproliferation and matrix remodelling; (3) immune dysregulation; and (4) vascular and endothelial damage. The aim is to review the rationale, potential and limitations of current candidate biomarkers and their utility in IPF and SSc-ILD to help direct future research and translation to clinical practice.