Plasma Cell-Free DNA Integrity Assessed by Automated Electrophoresis Predicts the Achievement of Pathologic Complete Response to Neoadjuvant Chemotherapy in Patients With Breast Cancer

Abstract

Purpose: The study of plasma cell-free DNA integrity (cfDI) has shown potential for providing useful information in neoplastic patients. The aim of this study is to estimate the accuracy of an electrophoresis-based method for cfDI evaluation in the assessment of pathologic complete response (pCR) in patients with breast cancer (BC) undergoing neoadjuvant chemotherapy (NACT).

Patients and methods: Fifty-one patients with BC undergoing anthracycline-/taxane-based NACT were recruited. Plasma samples were collected from each patient at diagnosis (t0), after anthracycline administration (t1), and after NACT completion (t2). The concentration of differently sized cell-free DNA fragments was assessed by automated electrophoresis. cfDI, expressed as cfDI index, was calculated as the ratio of 321-1,000 bp sized fragment concentration to 150-220 bp sized fragment concentration assessed at t2. cfDI index was then used to build an exploratory classifier for BC response to NACT, directly comparing its sensitivity and specificity with magnetic resonance imaging (MRI), through bootstrapped logistic regression.

Results: cfDI index was assessed on 38 plasma samples collected from as many patients at t2, maintaining a 30/70 ratio between pCR and non-pCR patients. cfDI index showed an area under the receiver operating characteristic curve in predicting the achievement of pCR of 81.6, with a cutoff above 2.71 showing sensitivity = 81.8 and specificity = 81.5. The combination of cfDI index and MRI showed, in case of concordance, an area under the receiver operating characteristic curve of 92.6 with a predictive value of complete response of 87.5 and a predictive value of absence of complete response of 94.7.

Conclusion: cfDI index measured after NACT completion shows great potential in the assessment of pCR in patients with BC. The evaluation of its use in combination with MRI is strongly warranted in prospective studies.

FIG 1.

cfDI index correlates with the response to neoadjuvant chemotherapy. (A) Boxplots of cell-free DNA fragment concentration assessed on 38 samples collected at t2 from as many patients (11 pCR and 26 nR). (B) Boxplot of 150-220 bp sized fragment concentration in six healthy controls, 27 nR, and 11 pCR patients at t2. The concentration of 150-220 bp sized fragments assessed at t2 was significantly higher in nR patients compared with the pCR ones (P = .048) and the healthy controls (P = .0096). No significant difference in the same parameter was observed between pCR patients and the healthy controls (P = .93). (C) Boxplot of cfDI index in nR and pCR patients. A significant correlation (P = .0014) between cfDI index and the achievement of pCR was observed. cfDI, cell-free DNA integrity; nR, noncomplete responders; pCR, pathologic complete response.

FIG 2.

cfDI index achieves a comparable accuracy to MRI in predicting pathologic complete response. (A) cfDI index ROC curve. AUC amounted to 0.816 (95% CI, 0.676 to 0.957). (B) MRI ROC curve. AUC amounted to 0.771 (95% CI, 0.614 to 0.928). AUC, area under the ROC curve; cfDI, cell-free DNA integrity; MRI, magnetic resonance imaging; ROC, receiver operating characteristic.

FIG 3.

The combination of MRI and cfDI index augments the accuracy in pathologic complete response prediction. MRI and cfDI index concordantly classified 27 of 38 patients with breast cancer. Of these, eight were classified as complete responders, whereas 19 as nR. MRI and cfDI index were discordant in 11 of 38 patients. Of these, five were classified as complete responders by MRI and as nR by cfDI index, whereas six were classified as nR by MRI and as complete responders by cfDI index. When concordant, cfDI index and MRI achieved a predictive value of complete response of 0.875 and a predictive value of noncomplete response of 0.947. cCR, clinical complete response; cfDI, cell-free DNA integrity; MRI, magnetic resonance imaging; nR, noncomplete responders; pCR, pathologic complete response.