Sensitivity and reliability of DNA-based mutation analysis by allele-specific digital PCR to follow resistant BCR-ABL1-positive cells
Abstract
Mutations in the kinase domain (KD) of BCR-ABL1 are known causes of resistance to tyrosine kinase inhibitor (TKI) treatment of chronic myeloid leukemia (CML) and Ph-positive acute lymphoblastic leukemia (Ph+ ALL), and detection is routinely performed at the transcript level. Sequencing remains the preferential technology and enables the identification of mutations at any position in the KD.
Next-generation sequencing (NGS) is nearly 1-log more sensitive than standard sequencing, thus significantly facilitating earlier detection of mutations [1,2,3]. Moreover, NGS recognizes the complexity of the mutation landscape and can distinguish between polyclonal and compound mutations, if certain conditions are followed [1, 4].
The most frequently used amplicon-based NGS approaches are based on sequencing of shorter overlapping PCR products covering the entire length of the KD, thus distant mutations that may co-occur within one cell cannot be reliably assessed. There is a possibility of sequencing of long-range PCR product of the whole KD of BCR-ABL1, however there were available limited approaches and capacities of NGS platforms [5].
The promising technology for reliable assessment of compound mutations in the KD of BCR-ABL1 are Oxford Nanopore technology or SMRT PacBio technology.