Analysis of PIK3CA mutations in the lysate of sentinel lymph nodes in patients with early breast cancer

Background Although one-step nucleic acid amplification (OSNA), which measures cytokeratin (CK) 19 mRNA copies, is used for intraoperative detection of sentinel lymph node (SN) metastasis, CK19 mRNA copy number may not always accurately reflect total tumor load in the SN. Because number of DNA copies per cell generally has smaller deviation, we hypothesized that detection of tumor-derived mutated DNA in SNs, by targeting genetic mutations in the primary tumor, may provide more accurate results than OSNA. We investigated the PIK3CA mutation, frequently detected in breast cancer, to explore the potential of this technique for diagnosing SN metastasis. Methods We analyzed data from 94 patients who had undergone SN biopsy at Osaka University Hospital (April 2017 to March 2019). Next-generation sequencing was used for mutation analysis of the primary tumor. In cases of PIK3CA mutations, OSNA lysates were analyzed to detect PIK3CA mutations in the SN by using droplet digital polymerase chain reaction (ddPCR). Results PIK3CA mutations were detected in 33.0% (31/94) of primary tumors, 25 of which had hotspot PIK3CA mutations and included 59 SNs. Of these SNs, 10 were diagnosed as metastasis positive by OSNA and confirmed by ddPCR to have PIK3CA mutations, with no false negatives. Conclusions Assessment of tumor-derived mutated DNA in SNs may be a useful technique to detect SN metastasis, as confirmed by ddPCR analysis. Further analyses, using data from a greater number of patients, are necessary to determine whether the results of whole-genome and whole-exome sequencing can be applied to other genes.