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Published on: March 29, 2017
Rapid and highly sensitive detection of MYD88/CXCR4 mutations in Waldenström macroglobulinemia by MAS-qPCR: A pilot
Yuki Kunimune1, Mitsuaki Nishioka2, Haruka Namba3
1Division of Laboratory, Yamaguchi University Hospital, Ube, Japan; Department of Oncology and Laboratory Medicine, Yamaguchi University Graduate School of Medicine, Ube, Japan.
Background:
Waldenström macroglobulinemia (WM) is a subset of lymphoplasmacytic lymphoma characterized by bone marrow infiltration of tumor cells and IgM monoclonal gammopathy. MYD88 mutations are found in more than 90% of WM patients and CXCR4 mutations in approximately 30-40%. For this reason, MYD88 mutation analysis is particularly important for the definitive diagnosis of WM. Furthermore, the CXCR4 S338* mutation is clinically significant for treatment selection in WM patients, as Bruton's tyrosine kinase inhibitors have been reported to be less effective in patients harboring both MYD88 and CXCR4 mutations. However, there are currently no simple methods available for the simultaneous detection of MYD88 and CXCR4 mutations.
Objective:
This study aimed to evaluate the multiplex allele-specific quantitative PCR (MAS-qPCR) assay we developed that is highly sensitive, inexpensive, and capable of simultaneously detecting these two WM-driver mutations using a single-tube reaction on a universal quantitative PCR instrument.
Method:
We established optimal PCR conditions using plasmid DNA containing MYD88 L265P(T > C) and CXCR4 S338*(C > G) or S338*(C > A) mutations. Using the established method, 12 patient specimens were used for comparison with existing methods.
Results:
The results demonstrated optimal PCR conditions at an annealing temperature of 64°C for 40 cycles. The detection sensitivity of MYD88 L265P(T > C), CXCR4 S338*(C > G), and CXCR4 S338*(C > A) mutations were 0.1% for all variants. The MAS-qPCR method successfully detected mutations in WM patient samples with high accuracy.
Conclusion:
MAS-qPCR may be a highly sensitive and cost-effective screening method for detecting MYD88 L265P and CXCR4 S338* mutations in WM.
