Cross-continent inter-laboratory transfer of a clinical mass spectrometry lectin magnetic bead assay (LeMBA-MS) for ovarian cancer serum glycoprotein biomarkers.

Abstract

e17586 Background: Accurate non-invasive detection of ovarian cancer can assist in pre-surgical triaging and screening for earlier diagnosis and better survival from ovarian cancer. Lectin magnetic bead-coupled mass spectrometry (LeMBA-MS) was used to discover ovarian cancer biomarkers using biobank samples from the UK and validated in a cohort from Australia (Dutt et al. 2023 Proteomics Clinical Applications 17(4):e2200114). This study evaluates LeMBA-MS as a clinical assay platform by determining its inter-laboratory transfer, robustness and reproducibility across reference serum and clinical samples. Methods: Proseek Bio prepared LeMBA processing reagents, which were shipped from Australia to Arcadia Life Sciences in Malaysia for this study. A dynamic multiple reaction monitoring (dMRM) method was transferred electronically. The study comprised three phases. Firstly, the dMRM method was implemented and optimized at Arcadia using a direct serum tryptic digest of a commercial serum. Then, the reference serum was processed using Proseek Bio reagent kit and reproducibility analyzed across 5 replicate runs, with 15% CV for 90% of the peptides as acceptance for reproducibility. Finally, ten Malaysian clinical serum samples (NMRR-18-544-40614), comprising five epithelial ovarian cancer and five benign cases, collected prior to treatment, were analyzed in triplicate. Results: All three phases of the study were successfully completed with minor adjustments and remote consultation. Two batches of lectin-beads were produced and post-shipment performance monitored using a standard protein showed no significant differences for all three peptides (t-test p > 0.18). The implemented dMRM method comprised 94 biomarkers and 6 control peptides. The pre-defined biomarker peptide reproducibility criteria were met for both the reference serum and the clinical samples: 85 biomarker peptides (90.4%) had CVs below 15%, while 9 (9.6%) exceeded 15%. The latter corresponded to low-abundance peptides, and will likely be removed from further development. Statistical analysis on the small independent dataset showed 28 peptides from 8 glycoproteins to be significantly different (adjusted p-value <0.05) between benign and epithelial ovarian cancer groups, with 25 peptides lower in cancer compared to benign. A principal component analysis plot showed clear separation of the groups. Conclusions: This inter-laboratory study demonstrates the robustness of LeMBA-MS as a suitable clinical assay format, and provides pilot independent clinical validation of candidate ovarian cancer biomarkers. Additional clinical validation studies in diverse populations are warranted to inform biomarker panel development.