Dose intensification strategy influences infliximab pharmacokinetics but not clinical response after the same number of doses.

BACKGROUND: The optimal infliximab dose intensification strategy to address secondary loss of response (LOR) remains unclear. This study aimed to compare clinical and pharmacokinetic outcomes following (i) upfront infliximab re-induction with (ii) ongoing 6-weekly dose interval shortening (DIS), after the same number of doses. METHODS: A prospective parallel cohort study of inflammatory bowel disease patients who required infliximab dose intensification for secondary LOR using (i) re-induction (i.e., repeat 5 mg/kg 0, 2, 6-week dosing) followed by 8-weekly maintenance or (ii) 6-weekly 5 mg/kg DIS was undertaken. Week 32 clinical response was the primary outcome, with secondary evaluation of infliximab pharmacokinetics and predictors of response. RESULTS: Of 104 patients, 54 underwent re-induction, and 50 underwent 6-weekly DIS; 43 per cohort had clinically active disease, with comparable baseline infliximab levels (2.03 vs 2.02 ug/mL, P = 0.83). Clinical response was similar across re-induction and DIS cohorts at weeks 12 (69.8 vs 65.1%) and 32 (53.5 vs 62.8%, each P > 0.50); however, both strategies demonstrated distinct pharmacokinetic profiles at weeks 6 (18.45 vs 5.36 ug/mL, P < 0.01), 12 (8.94 vs 5.96 ug/mL, P = 0.02) and 30 (3.89 vs 6.35 ug/mL, P = .0.02). In multivariable analyses, objectively verified active disease at baseline (OR 12.92, 95% CI [1.84-90.84], P = 0.01), subtherapeutic week 6 infliximab levels (OR 0.12, 95% CI [0.01, 0.99], P = 0.049) and week 12 clinical response (OR 5.44, 95% CI [1.20-19.97], P = 0.04) were associated with week 32 response, as were week 2 infliximab levels (OR 1.34, 95% CI [1.02-1.47], P = 0.04) following re-induction. Following re-induction, week 2 infliximab levels <15.6 ug/mL (AUROC 0.76, 95% CI [0.54-0.99], P < 0.05) predicted nonresponse at week 32. CONCLUSION: Dose intensification strategy impacted immediate and sustained infliximab levels but not clinical response. Upfront intensification was associated with short-term pharmacokinetic advantages, including predictors of response, that diminished with time. Hence, when applying upfront dose intensification, clinicians should consider continuing intensified dosing to sustain early pharmacokinetic advantages based on predictors of (non)response.

Serum protein electrophoresis and rheumatoid factor analysis is an effective screening strategy for cryoglobulinaemia.

Accurate serum cryoglobulin detection is important to allow prompt treatment but laboratory testing requires stringent pre-analytical conditions and has long turnaround times. Serum protein electrophoresis (EPG) for paraproteinaemia and rheumatoid factor (RF) analysis may offer an effective initial screening strategy for the presence of cryoglobulinaemia. We retrospectively assessed the sensitivity of ancillary EPG and RF testing for the presence of serum cryoglobulinaemia in 586 eligible cryoglobulin positive samples received at the Royal Prince Alfred and Liverpool Hospital immunopathology laboratories over an 11-year period. Ninety-one percent of all cryoglobulin positive samples had either a detectable paraprotein or RF activity, with greatest sensitivity for type I and type II cryoglobulins (97% and 98%, respectively). The sensitivity remained high irrespective of whether EPG and RF analysis was performed with the same, or different, pre-analytical collection conditions to the cryoglobulin collection (92% vs 90%, p=0.46). Only two patients with detected cryoglobulins and no associated paraprotein or RF activity had clinical features of cryoglobulinaemia and neither required treatment. This study demonstrates that serum EPG and RF analysis has high sensitivity for the detection of clinically relevant cryoglobulinaemia, even when not collected under ideal pre-analytical conditions, and potentially offers a prompt and effective screening strategy.

Timing of Live Attenuated Vaccination in Infants Exposed to Infliximab or Adalimumab in Utero: A Prospective Cohort Study in 107 Children.

BACKGROUND AND AIMS: For infants exposed in utero to anti-tumour necrosis factor-α [TNF] medications, it is advised that live-attenuated vaccinations be postponed until the drug is cleared, but little is known about time to clearance. To minimize delays before live-attenuated vaccination can be given, we aimed to develop a pharmacokinetic model to predict time-to-clearance in infants exposed during pregnancy. METHODS: We prospectively followed in utero infliximab/adalimumab-exposed infants of mothers with inflammatory bowel disease across four countries between 2011 and 2018. Infants with a detectable anti-TNF umbilical-cord level and at least one other blood sample during the first year of life were included. RESULTS: Overall, 107 infants were enrolled, including 166 blood samples from 71 infliximab-exposed infants and 77 samples from 36 adalimumab-exposed infants. Anti-TNF was detectable in 23% [n = 25] of infants at 6 months. At 12 months, adalimumab was not detected but 4% [n = 3] had detectable infliximab. A Bayesian forecasting method was developed using a one-compartment pharmacokinetic model. Model validation showed that the predicted clearing time was in accordance with the measured observations. A clinician-friendly online calculator was developed for calculating full anti-TNF clearing time: https://xiaozhu.shinyapps.io/antiTNFcalculator2/. CONCLUSIONS: Almost one-quarter of infants born to mothers receiving anti-TNF during pregnancy have detectable anti-TNF at 6 months. To limit the time to live-attenuated vaccination in infants of mothers receiving anti-TNF during pregnancy, the results of a cord drug level at birth and a second sample ≥ 1 month thereafter can be used to estimate the time for full anti-TNF clearance in these children.

Intrinsic Defects in B Cell Development and Differentiation, T Cell Exhaustion and Altered Unconventional T Cell Generation Characterize Human Adenosine Deaminase Type 2 Deficiency.

PURPOSE: Deficiency of adenosine deaminase type 2 (ADA2) (DADA2) is a rare inborn error of immunity caused by deleterious biallelic mutations in ADA2. Clinical manifestations are diverse, ranging from severe vasculopathy with lacunar strokes to immunodeficiency with viral infections, hypogammaglobulinemia and bone marrow failure. Limited data are available on the phenotype and function of leukocytes from DADA2 patients. The aim of this study was to perform in-depth immunophenotyping and functional analysis of the impact of DADA2 on human lymphocytes. METHODS: In-depth immunophenotyping and functional analyses were performed on ten patients with confirmed DADA2 and compared to heterozygous carriers of pathogenic ADA2 mutations and normal healthy controls. RESULTS: The median age of the patients was 10 years (mean 20.7 years, range 1-44 years). Four out of ten patients were on treatment with steroids and/or etanercept or other immunosuppressives. We confirmed a defect in terminal B cell differentiation in DADA2 and reveal a block in B cell development in the bone marrow at the pro-B to pre-B cell stage. We also show impaired differentiation of CD4+ and CD8+ memory T cells, accelerated exhaustion/senescence, and impaired survival and granzyme production by ADA2 deficient CD8+ T cells. Unconventional T cells (i.e. iNKT, MAIT, Vδ2+ γδT) were diminished whereas pro-inflammatory monocytes and CD56bright immature NK cells were increased. Expression of the IFN-induced lectin SIGLEC1 was increased on all monocyte subsets in DADA2 patients compared to healthy donors. Interestingly, the phenotype and function of lymphocytes from healthy heterozygous carriers were often intermediate to that of healthy donors and ADA2-deficient patients. CONCLUSION: Extended immunophenotyping in DADA2 patients shows a complex immunophenotype. Our findings provide insight into the cellular mechanisms underlying some of the complex and heterogenous clinical features of DADA2. More research is needed to design targeted therapy to prevent viral infections in these patients with excessive inflammation as the overarching phenotype.

Prospective randomised controlled trial of adults with perianal fistulising Crohn's disease and optimised therapeutic infliximab levels: PROACTIVE trial study protocol.

INTRODUCTION: Perianal fistulising Crohn's disease (pfCD) can be somewhat treatment refractory. Higher infliximab trough levels (TLIs) may improve fistula healing rates; however, it remains unclear whether escalating infliximab therapy to meet higher TLI targets using proactive, or routine, therapeutic drug monitoring (TDM) improves outcomes. This randomised controlled trial aimed to assess whether infliximab therapy targeting higher TLIs guided by proactive TDM improves outcomes compared with standard therapy. METHODS AND ANALYSIS: Patients with active pfCD will be randomised 1:1 to either the proactive TDM arm or standard dosing arm and followed up for 54 weeks. Patients in the proactive TDM arm will have infliximab dosing optimised to target higher TLIs. The targets will be TLI ≥ 25 µg/mL at week 2, ≥ 20 µg/mL at week 6 and ≥ 10 µg/mL during maintenance therapy. The primary objective will be fistula healing at week 32. Secondary objectives will include fistula healing, fistula closure, radiological fistula healing, patient-reported outcomes and economic costs up to 54 weeks. Patients in the standard dosing arm will receive conventional infliximab dosing not guided by TLIs (5 mg/kg at weeks 0, 2 and 6, and 5 mg/kg 8 weekly thereafter). Patients aged 18-80 years with pfCD with single or multiple externally draining complex perianal fistulas who are relatively naïve to infliximab treatment will be included. Patients with diverting ileostomies or colostomies and pregnant or breast feeding will be excluded. Fifty-eight patients per arm will be required to detect a 25% difference in the primary outcome measure, with 138 patients needed to account for an estimated 6.1% primary non-response rate and 10% dropout rate. ETHICS AND DISSEMINATION: Results will be presented in peer-reviewed journals and international conferences. Ethics approval has been granted by the South Western Sydney Local Health District Human Research Ethics Committee in Australia. TRIAL REGISTRATION NUMBER: Australian New Zealand Clinical Trials Registry (ACTRN12621000023853); Pre-results.

Therapeutic drug monitoring in inflammatory bowel disease reduces unnecessary use of infliximab with substantial associated cost-savings.

BACKGROUND: Therapeutic drug monitoring (TDM) of infliximab (IFX) levels in inflammatory bowel disease (IBD) patients can help to guide dose adjustments or changes to therapy for selected patients in remission or with secondary loss of response (LOR). AIMS: To determine how IFX TDM is utilised in a real-life clinical setting and to quantify the potential for TDM to reduce the unnecessary use of IFX. METHODS: Data from all public IBD IFX level testing performed across Australia were prospectively collected from June 2016 to July 2017 to assess physician-reported for testing indications (induction, in remission or LOR) and associated results. The hypothetical influence of IFX TDM was based on an optimal therapeutic range of 6-10 mg/L for mucosal healing. RESULTS: Secondary LOR (reactive TDM) was the most common indication for TDM. These patients have consistently lower median IFX levels: 3.02 mg/L (IQR 1.14-6.67 mg/L) versus 5.22 mg/L (IQR 2.70-8.12 mg/L), P = 0.0001 compared with patients in remission (proactive TDM). TDM helped to identify unnecessary use of IFX in 30.6% of the TDM tests performed in luminal Crohn disease and ulcerative colitis patients, with an associated drug cost saving of $531.38 per IFX TDM test episode. Unnecessary IFX use was identified in 38.9% (96/247) of reactive IFX TDM tests performed and in 19.3% (35/181) of proactive testing. CONCLUSION: Use of both reactive and proactive IFX TDM is cost-effective for IBD management as it informs the clinician where unnecessary use of IFX can be stopped.

Comparison of three commercially available ELISA assays for anti-infliximab antibodies.

Three commercially available assays for the measurement of antibodies to infliximab (ATI) are approved for clinical use in Australia: Promonitor anti-infliximab (Grifols), Lisa Tracker anti-infliximab (Theradiag) and Ridascreen anti-IFX (R-Biopharm). All are bridging ELISA assays. Measurement of ATI has been incorporated into treatment algorithms for assessing loss of response to infliximab in patients with inflammatory bowel disease, but results obtained by the three ATI assays have not been systematically compared. We performed a series of experiments to allow comparison of results between the assays. Forty-two patient samples known to be positive for ATI by the Lisa Tracker assay were run on the Promonitor assay in singlicate, of which 26 were run on the Ridascreen assay in duplicate, according to the manufacturers' instructions. The Spearman correlation coefficient for all three pairwise assay comparisons was 0.95. Results were not numerically comparable between the assays. The coefficient of variation (CV) was 2.3% for the Lisa Tracker assay, 7.6% for the Promonitor assay and 7.4% for the Ridascreen assay. The presence of infliximab interfered with all three assays in a dose dependent manner. The cut-point for loss of response to infliximab dose intensification, previously demonstrated to be 200 ng/mL on the Lisa Tracker assay, is equivalent to approximately 60 ng/mL on the Ridascreen assay and between 22.9 and 41 AU/mL on the Promonitor assay. All three assays are suitable for clinical use.

Comparative Evaluation of 4 Commercially Available ELISA Kits for Measuring Adalimumab and Anti-adalimumab Antibodies.

BACKGROUND: Therapeutic drug monitoring of tumor necrosis factor inhibitors, such as adalimumab (ADM), is increasingly being performed for the management of autoimmune diseases. However, there can be significant variation in drug and antibody concentrations obtained by different assay methods. The aim of this study was to compare the performance of 4 enzyme-linked immunosorbent assay (ELISA) kits for measuring ADM and anti-ADM antibodies. METHOD: Dilutions of ADM or anti-ADM spiked sera were assessed for recovery rate and precision using the following 4 kits: LISA-Tracker (Theradiag, Croissy-Beaubourg, France), Promonitor (Grifols, Barcelona, Spain), Ridascreen (R-Biopharm, Darmstadt, Germany), and Shikari (Matriks Biotek, Gölbasi/Ankara Turkey). Interference samples were also assessed. RESULTS: At the therapeutic concentration, ADM detection was comparable among the 4 ELISA kits. Lisa-Tracker and Shikari kits produced low-range false positive results in normal sera. Infliximab and etanercept caused false positives in Lisa-Tracker and Shikari kits. Anti-ADM antibody ELISA kits performed differently with spiked samples because of different measuring units and ranges. Ridascreen and Shikari kits were dose responsive across the entire standard curve and correlated well with each other (r = 0.997). Cross reactivity was observed in rheumatoid factor positive sera tested on the Promonitor anti-ADM kit. CONCLUSIONS: All ADM kits tested were dose responsive within the therapeutic range and correlated well. The significance of observed low-range false positives and cross reactivity with infliximab in LISA-Tracker and Shikari kits is dependent on the indications received for testing in the laboratory. Anti-ADM ELISA kits produced varied results for spiked sera; however, they showed good precision. Inter-kit variability suggested that anti-ADM levels should be compared only when using the same method.

Gastric Cancer Screening in Common Variable Immunodeficiency.

Individuals with common variable immunodeficiency (CVID) have an increased risk of gastric cancer, and gastrointestinal lymphoma, yet screening for premalignant gastric lesions is rarely offered routinely to these patients. Proposed screening protocols are not widely accepted and are based on gastric cancer risk factors that are not applicable to all CVID patients. Fifty-two CVID patients were recruited for screening gastroscopy irrespective of symptoms or blood results and were compared to 40 controls presenting for gastroscopy for other clinical indications. Overall, 34% of CVID patients had intestinal metaplasia (IM), atrophic gastritis or moderate to severe non-atrophic gastritis, which can increase the risk of gastric cancer, compared to 7.5% of controls (p < 0.01). Focal nodular lymphoid hyperplasia, a precursor lesion for gastrointestinal lymphoma, was seen in eight CVID patients (16%), one of whom was diagnosed with gastrointestinal lymphoma on the same endoscopy. High-risk gastric pathology was associated with increased time since diagnosis of CVID, smoking, Helicobacter pylori, a low-serum pepsinogen I concentration, and diarrhea, but not pepsinogen I/II ratio, iron studies, vitamin B12 levels or upper gastrointestinal symptoms. There was a lower rate of detection of IM when fewer biopsies were taken, and IM and gastric atrophy were rarely predicted by the endoscopist macroscopically, highlighting the need for standardized biopsy protocols. The prevalence of premalignant gastric lesions in patients with CVID highlights the need for routine gastric screening. We propose a novel gastric screening protocol to detect early premalignant lesions and reduce the risk of gastric cancer and gastric lymphoma in these patients.

Comparison of infliximab drug measurement across three commercially available ELISA kits.

The monitoring of infliximab drug levels aids in the management of several autoimmune diseases, notably inflammatory bowel disease. Several commercial kits are now available and approved by the Therapeutic Goods Administration (TGA) for the measurement of infliximab levels, but there have been limited verification or comparison studies to date. Finding an assay that most accurately measures infliximab is essential for optimal drug titration and patient management. We performed this study to compare the performance of the Grifols Promonitor, Theradiag Lisatracker and R-Biopharm Ridascreen enzyme linked immunosorbent assay (ELISA) kits. Preparations of serum containing known concentrations of infliximab were assayed using each kit, including in the presence of interference from anti-infliximab antibodies, autoantibodies and other biological agents. The Lisatracker kit provided the most accurate determination of infliximab drug levels, however it yielded false positive results at low concentrations of infliximab. The average coefficients of variation (CVs) for the kits were 8% for Lisatracker, 5% for Ridascreen and 11% for Grifols. Infliximab measurements across all kits were affected by interference from antibodies to infliximab (ATI). This study identified the Lisatracker kit as the most accurate in quantifying infliximab levels, although it was limited by false positive results at low concentrations of infliximab as well as interference from ATI. This has important implications for the monitoring and management of patients receiving infliximab therapy.

Anti-synthetase syndrome associated with anti PL-12 and anti-Signal recognition particle antibodies and a necrotizing auto-immune myositis.

We report a 37-year-old woman with a 2 month history of proximal muscle weakness and extremely high creatine kinase (21,808 U/L) due to necrotizing auto-immune myositis (NAM) in association with anti-synthetase syndrome. Myositis-specific auto-immune antibody panel was positive for anti-Signal recognition particle and anti-PL-12. CT scan of the chest confirmed interstitial lung disease. Prednisolone, intravenous immunoglobulin and cyclophosphamide therapy was given with gradual improvement. This patient is notable for the unusual combination of NAM and anti-synthetase syndrome with the rare finding of two myositis-specific autoantibodies, which directed testing for associated extramuscular features and management with more aggressive immunotherapy.

Use of cardiac MR imaging to evaluate the presence of myocarditis in autoimmune myositis: three cases.

Cardiac involvement in patients with idiopathic inflammatory myopathies (autoimmune myositis) is important to detect because it confers an increased risk of mortality. However, detection of myocardial involvement is hampered by a lack of sensitivity of traditional non-invasive methods, and the finding of elevated cardiac troponin T levels that may be due to regenerating skeletal muscle, rather than myocardial damage. Here, we describe three cases of inflammatory myositis with elevated troponin T levels, and non-specific echocardiographic and ECG findings. Cardiac MR imaging was useful in the evaluation for the presence of myocarditis or alternative cardiac pathology.

Clearing the complexity: immune complexes and their treatment in lupus nephritis.

Systemic lupus erythematosus (SLE) is a classic antibody-mediated systemic autoimmune disease characterised by the development of autoantibodies to ubiquitous self-antigens (such as antinuclear antibodies and antidouble-stranded DNA antibodies) and widespread deposition of immune complexes in affected tissues. Deposition of immune complexes in the kidney results in glomerular damage and occurs in all forms of lupus nephritis. The development of nephritis carries a poor prognosis and high risk of developing end-stage renal failure despite recent therapeutic advances. Here we review the role of DNA-anti-DNA immune complexes in the pathogenesis of lupus nephritis and possible new treatment strategies aimed at their control.