BCR-ABL1 (p210) Transcript Kinetics

Context.— Delta checks are a powerful technique for monitoring clinical assays in many disciplines but have not been routinely used in molecular testing. Objective.— To determine if the biologically determined kinetics of BCR-ABL1's rise and fall could allow the development of a delta check in BCR-ABL1 testing. Design.— Nine years of BCR-ABL1 p210 results were evaluated and patients with 3 or more results were selected for inclusion. The kinetics of these percentages of international standard values were plotted against time along with the median and the 90th and 95th percentile lines. A Monte Carlo simulation of a batch mix-up was performed for 6 months of data to determine the efficacy of the proposed cutoff. Results.— The median kinetics showed a 1-log drop of the percentage of international standard in 90 days, with less than 5% of cases showing faster than a 2-log drop in 90 days, and less than 2.5% showing a faster than 3-log drop in 90 days (extrapolated to 1 log in 30 days). The Monte Carlo simulation of a batch mix-up showed that an average batch mix-up of 23 samples could routinely be flagged by this cutoff, albeit with wide variance. Conclusions.— These results suggest that using a drop in the percentage of international standard of greater than 1 log in 30 days can be a useful trigger in implementing a delta-check system for this molecular test.

Analysis of MTb Rapid Molecular Test Performance Towards Microscopical Acid Fast Bacilli Examination at Labuang Baji General Hospital

Tuberculosis (TB) is a global health problem, which is the third leading cause of death of all infectious diseases aroundthe world, included Indonesia. Acid Fast Bacilli (AFB) smear and rapid molecular assay for Mycobacterium tuberculosis (MTB)are the old and new examinations required for MTB laboratory diagnosis. This study aimed to compare the performance ofMTB rapid molecular assay and AFB smear in diagnosis and screening for TB patients. This observational retrospective studyused a cross-sectional approach, with a purposive sampling technique of 559 patients with suspected TB in Labuang BajiHospital, Makassar. This study was conducted from March 2019 to June 2019 by taking data from medical records fromJanuary 2018 to December 2018 at Labuang Baji Hospital, Makassar. Three hundred and forty-nine subjects were males(62.4%), and 210 subjects were females (37.6%). This study revealed sensitivity and specificity of 98.57% and 84.96%,respectively for MTB rapid molecular assay, and 68.65% and 99.44%, respectively for AFB smear, this shows that MTB rapidmolecular assay was superior to AFB smear in diagnosing TB patients.

701 ECMO assistance during TAVI in high-risk patient with recent COVID-19 pneumonia

Methods and results A 87 years old woman, with history of dyslipidemia and permanent Atrial Fibrillation, already undergone full SARS-CoV2 vaccination few months before, referred to our E.R. with complain of dyspnoea and chest pain. COVID-19 molecular test resulted positive and CT Scan of the chest confirmed the presence of several areas of ground-glass opacity and consolidation together with bilateral pleural effusion (right 6 cm with pulmonary atelectasis; left 2 cm), not requiring drainage. Moreover, it showed severe calcification of both the aortic valve and root. Transthoracic echocardiogram showed eccentric LV hypertrophy with diffuse hypokinesia (EF 20–25%), ectatic ascending aorta (45 mm) with severe LF-LG aortic stenosis (AVAi 0.19 cm2) and moderate regurgitation, moderate-severe mitral regurgitation. During hospitalization in the COVID-19 Unit, despite O2 therapy she experienced worsening of the respiratory status with concomitant pulmonary oedema, hypotension and acute kidney injury, requiring administration of i.v. dobutamine and high dose diuretics. After gradual stabilization and COVID-19 negativization on 10th molecular test, she was transferred to our Coronary Care Unit. Coronary angiography showed absence of significant stenoses in the main vessels. In the following days the patient underwent a new clinical deterioration with dyspnoea, hypotension (BP 85/50 mmHg), oliguria and ankle swelling, requiring Ventimask O2 therapy and Dobutamine infusion. Transtoracic echocardiogram confirmed EF of 25% with PASP 30 mmHg. We decided to perform a ‘Rescue’ TAVI procedure, facilitated by extra-corporeal cardiac and respiratory support. CT Angiography of the chest, performed with low-dose contrast injection under amines infusion, showed severly calcific aortic valve with large sizes of the ring (Virtual Basal Ring area 620 mm2, perimeter 91 mm), measures compatible with the largest sizes of TAVI prostheses. After Veno-Arterial Extracorporeal Membrane Oxygenation (ECMO) cannulation, we performed the implantation of a 34 mm Evolut R (Medtronic) TAVI prosthesis, post-dilated with 24 mm balloon for under-expansion due to massive calcification. During both self-expandable TAVI delivery and balloon inflation the patient underwent two phases of cardiac arrest, during which the ECMO flow provided a proper circulatory support. Conclusions Since percutaneous valve replacement the patient’s recovery was fast with rapid ECMO removal and discontinuation of inotropic therapy. Few weeks after discharge, at first follow-up examination, the patient appeared asymptomatic, in excellent clinical conditions. 701 Figure

Results of the TCM Method of Pulmonary Tuberculosis Examination on Lung TB Suspects

Tuberculosis (TB) is an infectious disease caused by the bacterium Mycobacterium tuberculosis (MTB). Determinants of pulmonary TB disease include gender, and age. The Rapid Molecular Test Method (TCM) is a breakthrough TB program for Drug Resistant TB patients. The TCM method takes two hours to get the patient's diagnosis. TCM method for diagnosing TB and resistance to rifampin quickly and accurately. Tidar Magelang Hospital is one of the referral places for TCM examinations from various health facilities. The purpose of this study was to describe the results of the TCM method of pulmonary TB examination on pulmonary TB suspects at Tidar Magelang Hospital. This type of research is descriptive. The research data is secondary data obtained from Tidar Hospital Magelang. Based on age, namely early adolescence, late adolescence, early adulthood, late adulthood, early elderly, late elderly, and seniors, the results were sensitive rif 0.76%, 11.45%, 9, 54%, 7.25%, 5.73%, 8.78% and 3.82%. Negative results 1.15%, 1.91%, 6.87%, 7.25%, 7.63%, 14.89% and 12.98%. The rif results were sensitive to 27.1% males and 20.23% females. Negative results in 31.3% men and 21.37% women. The most sensitive rif results are in Magelang, followed by Magelang City, then East Jakarta Municipality, Banjarnegara and Purworejo. Negative results were found in Magelang, followed by Magelang City, then Wonosobo and East Waringin City.

Performance of the eazyplex® BloodScreen GN as a simple and rapid molecular test for identification of Gram-negative bacteria from positive blood cultures

The LAMP-based eazyplex® BloodScreen GN was evaluated for the detection of frequent Gram-negatives directly from positive blood culture (BC) bottles. A total of 449 BCs were analyzed. Sensitivities and specificities were 100% and 100% for Escherichia coli, 95.7% and 100% for Klebsiella pneumoniae, 100% and 100% for blaCTX-M, 100% and 100% for Klebsiella oxytoca, 100% and 99% for Proteus mirabilis, and 100% and 99.8% for Pseudomonas aeruginosa, respectively. The time to result ranged from 8 to 16 min, plus about 6 min for sample preparation. The eazyplex® BloodScreen GN is a reliable molecular assay for rapid BC testing.

Clinical impact of the rapid molecular detection of RSV and influenza A and B virus in the emergency room

Introduction: Using respiratory viruses' rapid diagnostic tests in the emergency room (ER) could allow a better and faster clinical management. Point-of-care PCR instruments provide now results in less than 30 minutes. The objective of this study was to assess the impact of the use of the cobas® Influenza A/B & RSV Assay for use on Roche's cobas® Liat® instrument on the clinical management of ER patients. Methods: Patients (adults and children) requiring admission or suffering from an underlying condition at risk of respiratory complication were prospectively recruited in the ER of four hospitals in the Brussels region. Physician's intentions regarding admission, isolation, antibiotic and antiviral use were collected before and after performing a cobas® Influenza A/B & RSV molecular test. Additionally, a comparison of the analytical performance of this test against antigen rapid tests and viral culture was performed as well as a time-to-result evaluation. Results: PCR yielded twice more positive results than antigen rapid diagnostic tests. It allowed a decrease in the overall need for isolation and treatment by limiting the isolation of negative patients and the antibiotic use for positive patients. Meanwhile, antiviral treatments better targeted patients with a positive influenza PCR. Conclusion: The use of a rapid influenza and RSV molecular test improves the clinical management of patients admitted to the ER by providing a fast and reliable result. Their additional cost compared to antigen tests should be balanced with the benefit of their analytical performance, leading to efficient reductions in the need of isolation and antibiotic use.

FINAL Analysis of a PAN European STOP Tyrosine Kinase Inhibitor Trial in Chronic Myeloid Leukemia : The EURO-SKI Study

Background: With the dramatic success of tyrosine kinase inhibitors (TKI) to treat Chronic myeloid leukemia (CML), the life expectancy of CML patients is now close to that of the general population. In addition, treatment cessation is now a realistic goal for some CML patients. This was shown by several clinical trials such as the STIM study leading to the concept of TFR (treatment free remission). Around 40-60% of patients with stable DMR [deep molecular response, corresponding to <0.01% BCR-ABL (IS)] can stop the TKI successfully, e.g. in accordance with recommendations from the European LeukemiaNet. In the interim analysis of the first 200 patients of the EURO-SKI (European Stop TKI) trial, 62% were in major molecular response (MMR: <0.1% BCR-ABL1 IS) at 6 months. DMR duration before TKI stop was most predictive for maintenance of MMR. Here we present the final analysis of the EURO-SKI trial after 3 years of follow-up. Aims: The main objectives of The EURO-SKI trial were the evaluation of molecular recurrence-free survival (MRecFS) after Stopping TKI in a large Pan-european cohort of CML patients and definition of prognostic markers to increase the rate of patients in durable deep MR after stopping TKI. Further aims are the evaluation of harmonized methods of molecular monitoring. Methods: Adult CML patients in chronic phase CML on TKI treatment in confirmed DMR for at least one year (confirmed by three consecutive PCR tests) and under TKI treatment for at least 3 years were eligible. DMR confirmation was performed in standardized laboratories. Primary endpoint was maintenance of MMR after stopping TKI. According to protocol, a 36 months follow-up was planned. The null hypotheses were that MMR maintenance at 6 and 36 months was less or equal than 40% and less or equal than 35%, respectively. Results: Between May 2012 and December 2014, 868 patients were pre-registered by 61 centers from 11 countries. 140 pts were excluded (consent withdrawal n=1, protocol violation n=38, not eligible n=74, DMR not confirmed n=11, atypical/unknown transcript n=15, missing data n=1) resulting in 728 eligible patents. Of these, 46.8% were female. Median age at diagnosis was 52 years (range, 11 to 85 years). Median duration of TKI treatment was 7.5 years (range, 3.0-14.1 years) and median duration of MR4 before TKI cessation was 4.7 years (range, 1.0-13.3 years). Nine patients died without MMR loss (none CML related), 15 patients restarted TKI without MMR loss. At 6 months, 713 patients were available (without molecular test at 6 months: n=6, TKI restart without relapse: n=9). Since 434 patients (61%) [95% CI: 57-64] remained without relapse during the first 6 months, the null hypothesis was rejected (p<0.0001). At 36 months, 678 patients could be analyzed (TKI restart without relapse: n=17, no molecular test at 36 months: n=33). With 309 patients in MMR, corresponding to 46% [95% CI: 42-49], the null hypothesis of 35% or less was rejected (p<0.0001). MRecFS at 36 months resulted in 48% (CI: 44-52%) and molecular recurrence- and treatment-free survival (MRecTFS) in 46% (CI: 43-50%) (Fig 1). No blast crisis occurred. Regarding prognostic factors, we confirmed that TKI treatment duration and DMR duration were still the most important factors to predict MMR loss at 6 months. For the late recurrence, i.e., between 6 and 36 months (57 patients), TKI treatment duration before stop was the only relevant variable in a preliminary univariate logistic analysis. Summary/Conclusion: With this final analysis of the largest TFR trial, we confirm the MRecFS and MRecTFS rates at 6 months previously obtained from the interim analysis. However, late molecular recurrence (15% between 6 and 36 months) occurred and the underlying mechanisms need to be discussed. Nevertheless, 46% of the patients, were still in MRecTFS at 3 years. Figure 1 Figure 1. Disclosures Hochhaus: Bristol-Myers Squibb: Research Funding; Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding; Pfizer: Research Funding; Incyte: Research Funding. Hjorth-Hansen: AOP: Research Funding; Novartis: Research Funding; Pfizer: Research Funding. Mustjoki: Novartis: Research Funding; BMS: Research Funding; Janpix: Research Funding; Pfizer: Research Funding. Martínez-López: Roche, Novartis, Incyte, Astellas, BMS: Research Funding; Janssen, BMS, Novartis, Incyte, Roche, GSK, Pfizer: Consultancy. Nicolini: Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: travel, accommodations, expenses, Research Funding; Incyte Biosciences: Honoraria, Other: travel, accommodations, expenses, Research Funding, Speakers Bureau; Kartos Therapeutics: Consultancy, Membership on an entity's Board of Directors or advisory committees; Sun Pharma Ltd.: Consultancy, Membership on an entity's Board of Directors or advisory committees; BMS: Honoraria. Saussele: Roche: Honoraria; Pfizer: Honoraria; Incyte: Honoraria, Research Funding; BMS: Honoraria, Research Funding; Novartis: Honoraria, Research Funding.

495. Evaluation of Antigen Testing for Detection of COVID-19 Vaccine Breakthrough Cases in Long-Term Care

Background Long-term care facilities (LTCFs) are at high risk for severe COVID-19 outbreaks due to their congregate nature and vulnerable population. Oregon Health Authority (OHA) deployed point-of-care antigen (Ag) tests to promptly identify COVID-19 cases in LTCFs. However, their performance in identifying vaccine breakthrough cases has not been evaluated. Methods During 2/25/21–5/25/21, OHA supported testing of residents and staff for two outbreaks at a single LTCF. Paired nasal swabs were collected and tested for SARS-CoV-2 by CDC Influenza SARS-CoV-2 Multiplex PCR Assay (molecular test) and Abbott BinaxNOW COVID-19 Ag Card (Ag test) twice weekly during the outbreaks. Participants were considered fully vaccinated if ≥ 14 days had passed since completion of a vaccine series; all others were deemed unvaccinated. A vaccine breakthrough case was defined as a positive Ag or molecular test from a fully vaccinated person’s specimen. Performance characteristics of the Ag test were assessed, with molecular test as the reference standard. Cycle threshold (Ct) values were compared by one-sided independent t-tests. Results 94 unvaccinated residents and staff provided 563 paired samples; SARS-CoV-2 was detected in 21 (12 by Ag and molecular test, 6 by molecular test only, 3 by Ag test only), yielding Ag test sensitivity of 66.7% (95% CI: 43.8–83.7%) and specificity of 99.4% (95% CI: 98.4–99.8%). Mean Ct values were higher for specimens positive by PCR but negative by Ag than those positive by both (30.0 vs. 20.7, P < .01). 81 vaccinated persons provided 925 paired samples; SARS-CoV-2 was detected in 5 (1 by Ag and molecular test, 4 by molecular test only), yielding Ag test sensitivity of 20% (95% CI: 3.6–62.5%) and specificity of 100% (95% CI: 99.6–100%). Mean Ct values for specimens from vaccinated cases were higher than those from unvaccinated cases (30.2 vs. 23.8, P < .05). The lone Ag-positive breakthrough case had a Ct of 20; all others had Ct > 29. Conclusion Ag test performance and reduced sensitivity on specimens with high Ct values found in this population are consistent with published data. Molecular testing maximizes identification of vaccine breakthrough cases. More studies are needed to estimate the proportion of breakthrough cases missed by Ag testing and their risk of transmitting the virus in LTCFs. Disclosures All Authors: No reported disclosures

Clinical performance of the Roche Elecsys SARS-CoV-2 Antigen fully automated electrochemiluminescence immunoassay

Objective: We assessed the clinical performance of novel Roche Elecsys SARS-CoV-2 Antigen fully automated electrochemiluminescence immunoassay (ECLIA). Design and Methods: We tested 160 subjects, 110 (68.8%) with positive molecular test for SARS-CoV-2 infection in nasopharyngeal samples with Altona Diagnostics RealStar SARS-CoV-2 RT-PCR Kit and Roche Elecsys SARS-CoV-2 Antigen. The local imprecision was validated by analyzing three nasopharyngeal samples with different antigen concentration (1.84, 9.51 and 423.30 TCID50/mL) for 20 consecutive times (intra-assay imprecision) or for 10 consecutive working days (inter-assay imprecision).Results: The local intra- and inter-assay imprecision of Elecsys SARS-CoV-2 Antigen ECLIA was found to be comprised between 2.0-2.0% and 5.8-7.6%, yielding to a total imprecision of 6.2-7.8%. Highly significant correlation was found between Elecsys SARS-CoV-2 Antigen ECLIA and cycle threshold (Ct) values of SARS-CoV-2 S and E genes (both r=-0.91; p<0.001). The area under the curve (AUC), sensitivity and specificity of Elecsys SARS-CoV-2 Antigen ECLIA were 0.83, 0.43 and 1.00 in all samples, 0.99, 0.87 and 0.99 in those with both Ct values <30, as well as 1.00, 1.00 and 0.89 in samples with both Ct values <25. Conclusion: Roche Elecsys SARS-CoV-2 Antigen ECLIA may be a surrogate of molecular testing for identification of super-spreaders.