Serial monitoring of plasma DNA mutations in estrogen receptor positive metastatic breast cancer (ER + MBC) holds promise as an early predictor of therapeutic response. Here, we developed dPCR-SEQ, a customized assay that utilizes digital PCR-based target enrichment followed by next-generation sequencing to analyze plasma DNA mutations in ESR1, PIK3CA, and TP53. We validated dPCR-SEQ in a prospective cohort of 58 patients with ER + MBC and demonstrate excellent concordance with hotspot ESR1 mutation abundance measured by conventional digital PCR. The dPCR-SEQ assay revealed ESR1, PIK3CA, and TP53 plasma ctDNA mutations in 55%, 32%, and 32% of the study patients, respectively. We also observed dynamic changes in ESR1, PIK3CA, and TP53 ctDNA mutant allele fraction (MAF) that were frequently discordant between the different genes. Thus, monitoring plasma DNA mutation dynamics using a dPCR-SEQ assay is feasible, accurate, and may be investigated as a biomarker of therapeutic response in ER + MBC.

对雌激素受体阳性转移性乳腺癌 (ER + MBC) 血浆 DNA 突变的连续监测有望成为治疗反应的早期预测因子。在这里，我们开发了 dPCR-SEQ，这是一种定制的检测方法，利用基于数字 PCR 的靶标富集，然后进行新一代测序来分析ESR1、PIK3CA和TP53中的血浆 DNA 突变。我们在 58 名 ER + MBC 患者的前瞻性队列中验证了 dPCR-SEQ，并证明与传统数字 PCR 测量的热点ESR1突变丰度具有极好的一致性。dPCR-SEQ 测定分别显示 55%、32% 和 32% 的研究患者存在ESR1、PIK3CA和TP53血浆 ctDNA 突变。我们还观察到ESR1、PIK3CA和TP53 ctDNA 突变等位基因分数 (MAF) 的动态变化，这些变化在不同基因之间经常不一致。因此，使用 dPCR-SEQ 测定监测血浆 DNA 突变动态是可行的、准确的，并且可以作为 ER + MBC 治疗反应的生物标志物进行研究。