No abstract

Tyrosine kinase inhibitors (TKIs) targeting the BCR-ABL1 fusion protein, encoded by the Philadelphia chromosome, have drastically improved the outcomes for patients with chronic myeloid leukemia (CML). Although several real-time quantitative polymerase chain reaction (RQ-PCR) kits for the detection of BCR-ABL1 transcripts are commercially available, their accuracy and efficiency in laboratory practice require reevaluation. We have developed a new in-house RQ-PCR method to detect minimal residual disease (MRD) in CML cases. MRD was analyzed in 102 patients with CML from the DOMEST study, a clinical trial to study the rationale for imatinib mesylate discontinuation in Japan. The BCR-ABL1 / ABL1 ratio was evaluated using the international standard (IS) ratio, where IS < 0.1% was defined as a major molecular response. At enrollment, BCR-ABL1 transcripts were undetectable in all samples using a widely-applied RQ-PCR method performed in the commercial laboratory, BML (BML Inc., Tokyo, Japan); however, the in-house method detected the BCR-ABL1 transcripts in five samples (5%) (mean IS ratio: 0.0062 ± 0.0010%). After discontinuation of imatinib, BCR-ABL1 transcripts were detected using the in-house RQ-PCR in 21 patients (21%) that were not positive using the BML method. Nineteen samples were also tested using a commercially available RQ-PCR assay kit with a detection limit of IS ratio, 0.0032 (ODK-1201, Otsuka Pharmaceutical Co., Tokyo, Japan). This method detected low levels of BCR-ABL1 transcripts in 14 samples (74%), but scored negative for five samples (26%) that were positive using the in-house method. From the perspective of the in-house RQ-PCR method, number of patients confirmed loss of MMR was 4. These data suggest that our new in-house RQ-PCR method is effective for monitoring MRD in CML.

Clinical demand for the prompt assessment of the activity of direct-acting factor Xa inhibitors in the emergency care setting is increasing. In the present study, we examined whether prothrombin time (PT) tests can serve as a clinically useful indicator of anti-factor Xa activity. In the first series, the in vitro effect of edoxaban on PT was evaluated by spiking human plasma with edoxaban and measuring PT using three different commercial PT tests. In the second series, the reversal effect of prothrombin complex concentrates (PCC) and activated PCC (aPCC) in edoxaban-spiked plasma was evaluated. In the third series, PT of plasma samples from patients administered either 15 or 30 mg/day of edoxaban was assessed, and the results were compared with edoxaban concentrations determined by a calibrated anti-factor Xa activity assay. The spike test revealed that all PT reagents positively correlated with edoxaban. The sensitivity to edoxaban varied among the three reagents and Triniclot(®) Excel S showed the best performance. Prolonged PT by edoxaban was reversed by PCC and aPCC in a dose-dependent manner; however, complete reversal was not achieved. Positive correlation between anti-factor Xa activity and PT was shown in the clinical samples at the edoxaban range from 0 to >300 ng/mL.

Fondaparinux (FPX), a synthesized factor Xa inhibitor, is one of the most popular anticoagulants for the prevention of postoperative venous thromboembolism (VTE). Although routine monitoring is not required, the bleeding adverse events cannot be neglected, and the measurement of anti-Xa activity is expected to be monitored. The primary purpose of this study is to evaluate the performances of 2 chromogenic assays for the detection of anti-Xa activity. Furthermore, the pharmacokinetics of FPX was examined using chromogenic assays. Anti-Xa activity was measured using 2 FPX-based chromogenic substrates (S2222 and STA-Liquid Anti-Xa). The reproducibility, detection limits, linearity, and correlations between the substrates were examined using normal plasma doped with low and high concentrations of FPX formulation. In addition, anti-Xa activity in 235 clinical samples from 164 cases treated was measured, and the pharmacokinetics of FPX was evaluated. Both of the tested substrates were capable of accurately measuring the anti-Xa activity of FPX, with a lower limit of 0.05 μg/mL and a coefficient of variation of less than 10%. The repeated administration of FPX induced a gradual but significant increase in the anti-Xa activity, which was negatively correlated with body weight and estimated glomerular filtration rate. No significant correlation between the anti-Xa activity and the occurrence of postoperative VTE or bleeding event was observed. Anti-Xa activity can be successfully determined using 2 chromogenic assays and automated biochemical analyzers. The clinical significance of anti-Xa activity monitoring should be examined in the future study.

COVID-19 antibody testing has been developed to investigate humoral immune response in SARS-CoV-2 infection. To assess the serological dynamics and neutralizing potency following SARS-CoV-2 infection, we investigated the neutralizing (NT) antibody, anti-spike, and anti-nucleocapsid antibodies responses using a total of 168 samples obtained from 68 SARS-CoV-2 infected patients. Antibodies were measured using an authentic virus neutralization assay, the high-throughput laboratory measurements of the Abbott Alinity quantitative anti-spike receptor-binding domain IgG (S-IgG), semiquantitative anti-spike IgM (S-IgM), and anti-nucleocapsid IgG (N-IgG) assays. The quantitative measurement of S-IgG antibodies was well correlated with the neutralizing activity detected by the neutralization assay (r = 0.8943, p < 0.0001). However, the kinetics of the SARS-CoV-2 NT antibody in severe cases were slower than that of anti-S and anti-N specific antibodies. These findings indicate a limitation of using the S-IgG antibody titer, detected by the chemiluminescent immunoassay, as a direct quantitative marker of neutralizing activity capacity. Antibody testing should be carefully interpreted when utilized as a marker for serological responses to facilitate diagnostic, therapeutic, and prophylactic interventions.