Minimal residual disease detected by droplet digital PCR in peripheral blood stem cell grafts has a prognostic impact on high-risk neuroblastoma patients

Abstract

More than half of high-risk neuroblastoma (NB) patients have experienced relapse due to the activation of chemoresistant minimal residual disease (MRD) even though they are treated by high-dose chemotherapy with autologous peripheral blood stem cell (PBSC) transplantation. Although MRD in high-risk NB patients can be evaluated by quantitative PCR with several sets of neuroblastoma-associated mRNAs (NB-mRNAs), the prognostic significance of MRD in PBSC grafts (PBSC-MRD) is unclear. In the present study, we collected 20 PBSC grafts from 20 high-risk NB patients and evaluated PBSC-MRD detected by droplet digital PCR (ddPCR) with 7NB-mRNAs (CRMP1, DBH, DDC, GAP43, ISL1, PHOX2B, and TH mRNA). PBSC-MRD in 11 relapsed patients was significantly higher than that in 9 non-relapsed patients. Patients with a higher PBSC-MRD had a lower 3-year event-free survival (P = 0.0148). The present study suggests that PBSC-MRD detected by ddPCR with 7NB-mRNAs has a prognostic impact on high-risk NB patients.

Keywords: Droplet digital PCR (ddPCR); Minimal residual disease (MRD); Neuroblastoma (NB); Neuroblastoma-associated mRNAs (NB-mRNAs); Peripheral blood stem cell (PBSC).

Figure 1

Clinical course of high-risk NB patients. All patients were subjected to PBSC harvest after three or four cycles of IC and PBSC infusion after HDC. 10 and 1 patients received RA and GD2 for maintenance therapy, respectively. Among 20 patients, 11 patients relapsed and were dead. IC, induction chemotherapy; HDC, high-dose chemotherapy; RT, radiation therapy, RA, 13-cis-retinoic acid; GD2, anti-GD2 antibodies.

Figure 2

PBSC-MRD in high-risk NB patients. The levels of each NB-mRNA (CRMP1, DBH, DDC, GAP43, ISL1, PHOX2B, and TH mRNA) (a) and 7NB-mRNAs (b) in 20 PBSC grafts were determined by ddPCR.

Figure 3

Association between the level of 7NB-mRNAs in PBSC grafts and relapse of high-risk NB patients. (a) The level of 7NB-mRNAs was compared between 9 non-relapsed and 11 relapsed PBSC grafts. (b) The ROC curve was plotted for the level of 7NB-mRNAs in 9 non-relapsed and 11 relapsed PBSC grafts. ROC, the receiver operator characteristic; AUC, the area under curve; TV, threshold value.

Figure 4

Kaplan-Meier curve analysis for high-risk NB patients dichotomized by PBSC-MRD. Kaplan-Meier curve was plotted for EFS (a) and OS (b) of high-risk NB patients with high- and low-PBSC-MRD. EFS, event-free survival; OS, overall survival.