Background: Acute myeloid leukemia (AML) is a hematological malignancy characterized by genomic heterogeneity throughout disease progression and between patients. Traditionally, both multiparameter flow cytometry (MFC) and next-generation sequencing (NGS) samples are collected via bone marrow (BM) aspiration, which is an invasive procedure with risk of infection. Peripheral blood (PB) has been demonstrated as a good substitute for BM but the concordance at different levels of NGS variant allele frequency (VAF) has not been defined. This study aims to demonstrate the relationship of NGS signals between PB and BM.

Method: Genomic profiles of 60 matched samples of BM and PB were obtained retrospectively from 24 patients with de novo AML (n=17) or secondary AML (n=7). The samples were subjected to library preparation with a hematologic panel based on AmpliMark, an ultrasensitive amplicon-based NGS platform technology. The hematologic panel covers up to 45 genes common in myeloid and lymphoid neoplasms, with an established limit of detection of 0.1% VAF. Optimized bioinformatics pipeline and in-house sequencing noise removal algorithm were applied for variant selection. One of the secondary AML patients had a concurrent diagnosis of multiple myeloma; for the purposes of this analysis, the myeloma-associated variants were excluded.

Results: 50 out of 60 matched BM and PB samples (83.3%) in the cohort were positive for at least one variant in either sample type. Complete or partial concordance of genetic profiles was achieved in 86.7% of the matched BM and PB samples. All samples obtained during diagnosis or relapse showed complete concordance, while all partial concordant and discordant profiles were sampled during post-treatment or remission stages. All BM samples with ≥1.5% abnormal cells detected by MFC demonstrated complete concordance between BM and PB genetic profiles. Analysis at the individual variant level (n=115) showed 78.6% of positive percent agreement (PPA) comparing PB to BM in variant detection. For variants detected at ≥0.5% and ≥0.1% VAF in the BM samples, 100% PPA and 86.4% PPA were achieved from matched PB samples, respectively. A strong correlation of VAF (r=0.98) was also demonstrated between variants detected in BM and PB.

Conclusion: PB NGS is particularly sensitive for detection of myeloid specific mutations in de novo and secondary AMLs with complete concordance at VAFs ≥0.5%, when compared to BM. The high concordance and minimally invasive nature of PB sampling makes it a favorable option for diagnostic and monitoring of AML.

Citation Format: Benjamin Lim, Chae Yin Cher, Jonathan Poh, Kao Chin Ngeow, Zi Yi Lim, Min-Han Tan. Complete mutation concordance with amplicon-based NGS between peripheral blood and bone marrow at 0.5% VAF threshold [abstract]. In: Proceedings of the AACR Special Conference: Acute Myeloid Leukemia and Myelodysplastic Syndrome; 2023 Jan 23-25; Austin, TX. Philadelphia (PA): AACR; Blood Cancer Discov 2023;4(3_Suppl):Abstract nr A22.