Rapid and Reproducible Karyotyping with Long Read Sequencing in AML Patients
Michael Heuser, Anna Dolnik, Isabell Arnhardt, Courteney K Lai, Ekaterina Jahn, Mustafa Salim, Gudrun Göhring, Yvonne Behrens, Jonathan L. Lühmann, Doris Steinemann, Martin Neugebohren, Jens-Florian F Schrezenmeier, Yasmine Alwie, Arnold Kloos, Gesa Baurmann, Jörg Westermann, Frederik Damm, Hartmut Döhner, Arnold Ganser, Felicitas R Thol, Olga Blau, Konstanze Döhner, Lars Bullinger, Razif Gabdoulline, Jan Eric SträngAcute myeloid leukemia (AML) is characterized by recurrent chromosomal abnormalities that form the basis of the European LeukemiaNet (ELN) risk classification and serve as essential determinants of prognosis and therapeutic decision-making. Conventional metaphase karyotyping remains the diagnostic gold standard for detecting these abnormalities; however, its utility is limited by longer turnaround times, often delaying critical clinical management. Here, we present a long-read sequencing-based (LRS) low-coverage whole genome sequencing (lcWGS) approach using Oxford Nanopore Technology as a rapid and scalable alternative for cytogenetic profiling. A total of 100 diagnostic AML samples were analyzed, comprising 50 retrospectively selected cases with known adverse-risk cytogenetics and 50 prospectively enrolled patients with clinically defined de novo AML. LcWGS demonstrated robust analytical performance, identifying chromosomal aberrations with 93% sensitivity, specificity, and overall accuracy, respectively. Complex karyotypes were reliably detected, with an area under the curve (AUC) of 0.971. Reproducibility was validated through replicate sequencing at two independent laboratories (R=0.99). LcWGS-derived estimates of clone size showed moderate correlation with conventional cytogenetic assessments (R=0.54). Patients with complex karyotypes identified by lcWGS exhibited significantly shorter overall and relapse-free survival, closely mirroring outcomes defined by conventional karyotyping and underscoring the value of lcWGS for risk stratification. Median turnaround time from sample receipt to bioinformatics interpretation was approximately 34 hours, enabling delivery of actionable karyotype results within 72 hours. These findings establish lcWGS as a rapid, reproducible, and accurate platform for detecting clinically relevant chromosomal abnormalities, addressing a critical need for timely risk stratification and treatment initiation in AML.