Bone marrow involvement and subclonal diversity impairs detection of mutated CXCR4 by diagnostic next‐generation sequencing in Waldenström macroglobulinaemia

Summary CXCR4 mutations impact disease presentation and treatment outcomes in Waldenström macroglobulinaemia (WM). Non‐uniform testing for CXCR4 mutations may account for discordant findings in WM clinical trials. We compared two approaches used in these trials for detection of the most common CXCR4 (S338X) variant: targeted next‐generation sequencing (NGS) using unselected bone marrow (BM) samples, and combined allele‐specific polymerase chain reaction (AS‐PCR) and Sanger sequencing with unselected and CD19‐selected BM samples. Our findings showed that targeted NGS frequently yielded false‐negative results. Both CD19 selection and AS‐PCR markedly improved detection of CXCR4S338X mutations. Sensitivity was adversely impacted by low BM involvement and CXCR4 mutation clonality.