Level of Seven Neuroblastoma-Associated mRNAs Detected by Droplet Digital PCR Is Associated with Tumor Relapse/Regrowth of High-Risk Neuroblastoma Patients

Abstract

Monitoring of several sets of neuroblastoma-associated mRNAs (NB-mRNAs) by real-time quantitative PCR (qPCR) can be used to evaluate minimal residual disease in NB patients. Droplet digital PCR (ddPCR) is an adaption of qPCR that potentially provides simpler and more reproducible detection of low levels of mRNAs. However, whether minimal residual disease in NB patients can be monitored by ddPCR using a set of NB-mRNAs is not yet tested. In this study, 208 bone marrow (BM) and 67 peripheral blood samples were retrospectively collected from 20 high-risk NB patients with clinical disease evaluation at two Japanese centers between 2011 and 2018, and level of each NB-mRNA (CRMP1, DBH, DDC, GAP43, ISL1, PHOX2B, and TH mRNAs) was determined by ddPCR. Level of 7NB-mRNAs (defined as the combined signature of each NB-mRNA) was higher in BM than peripheral blood, but correlated significantly with each other. In accordance with disease burden, it varied with disease status (remission, stable, or progression) and collection time point (diagnosis, treatment, post-treatment, or relapse). In 73 post-treatment BM samples, it was significantly higher in 17 relapsed/regrown samples than in 56 nonrelapsed/nonregrown samples. Furthermore, ddPCR had a better prognostic value than qPCR in detecting 7NB-mRNAs in the same 73 post-treatment BM samples. This study suggests that ddPCR detection of 7NB-mRNAs is significantly associated with tumor relapse/regrowth in high-risk NB patients. Monitoring of several sets of neuroblastoma-associated mRNAs (NB-mRNAs) by real-time quantitative PCR (qPCR) can be used to evaluate minimal residual disease in NB patients. Droplet digital PCR (ddPCR) is an adaption of qPCR that potentially provides simpler and more reproducible detection of low levels of mRNAs. However, whether minimal residual disease in NB patients can be monitored by ddPCR using a set of NB-mRNAs is not yet tested. In this study, 208 bone marrow (BM) and 67 peripheral blood samples were retrospectively collected from 20 high-risk NB patients with clinical disease evaluation at two Japanese centers between 2011 and 2018, and level of each NB-mRNA (CRMP1, DBH, DDC, GAP43, ISL1, PHOX2B, and TH mRNAs) was determined by ddPCR. Level of 7NB-mRNAs (defined as the combined signature of each NB-mRNA) was higher in BM than peripheral blood, but correlated significantly with each other. In accordance with disease burden, it varied with disease status (remission, stable, or progression) and collection time point (diagnosis, treatment, post-treatment, or relapse). In 73 post-treatment BM samples, it was significantly higher in 17 relapsed/regrown samples than in 56 nonrelapsed/nonregrown samples. Furthermore, ddPCR had a better prognostic value than qPCR in detecting 7NB-mRNAs in the same 73 post-treatment BM samples. This study suggests that ddPCR detection of 7NB-mRNAs is significantly associated with tumor relapse/regrowth in high-risk NB patients. Neuroblastoma (NB) is the most common extracranial solid tumor in children and is characterized by its extreme heterogeneity, ranging from spontaneous regression to malignant progression.1Brodeur G.M. Neuroblastoma: biological insights into a clinical enigma.Nat Rev Cancer. 2003; 3: 203-216Crossref PubMed Scopus (1572) Google Scholar,2Nakagawara A. Li Y. Izumi H. Muramori K. Inada H. Nishi M. Neuroblastoma.Jpn J Clin Oncol. 2018; 48: 214-241Crossref PubMed Scopus (53) Google Scholar Approximately half of the patients with NB have high-risk disease, whose long-term survival remains as low as 40%. Although intensive multimodal treatment including high-dose chemotherapy with autologous stem cell rescue, surgery, radiotherapy, and immunotherapy have been introduced over the past decades, 50% of high-risk patients still experience tumor relapse/regrowth caused by chemoresistant minimal residual disease (MRD).3Maris J.M. Hogarty M.D. Bagatell R. Cohn S.L. Neuroblastoma.Lancet. 2007; 369: 2106-2120Abstract Full Text Full Text PDF PubMed Scopus (1492) Google Scholar, 4Maris J.M. Recent advances in neuroblastoma.N Engl J Med. 2010; 362: 2202-2211Crossref PubMed Google Scholar, 5Tolbert V.P. Matthay K.K. Neuroblastoma: clinical and biological approach to risk stratification and treatment.Cell Tissue Res. 2018; 372: 195-209Crossref PubMed Scopus (72) Google Scholar MRD is defined conceptually as residual tumor cells that persistently reside in cancer patients after local and systemic cancer therapies.6Burchill S.A. Selby P.J. Molecular detection of low-level disease in patients with cancer.J Pathol. 2000; 190: 6-14Crossref PubMed Scopus (48) Google Scholar It persists as cancer stem cells in primary tumor, circulating tumor cells in peripheral blood (PB), disseminated tumor cells in bone marrow (BM) and lymph nodes, and micrometastasis in other tissues. These cancer stem cells, circulating tumor cells, and disseminated tumor cells represent the dynamics of MRD in cancer patients.7Mordant P. Loriot Y. Lahon B. Castier Y. Lesèche G. Soria J.-C. Massard C. Deutsch E. Minimal residual disease in solid neoplasia: new frontier or red-herring?.Cancer Treat Rev. 2012; 38: 101-110Abstract Full Text Full Text PDF PubMed Scopus (13) Google Scholar,8Uemura S. Ishida T. Thwin K.K.M. Yamamoto N. Tamura A. Kishimoto K. Hasegawa D. Kosaka Y. Nino N. Lin K.S. Takafuji S. Mori T. Iijima K. Nishimura N. Dynamics of minimal residual disease in neuroblastoma patients.Front Oncol. 2019; 9: 455Crossref PubMed Scopus (10) Google Scholar Because of the invasive nature of surgical biopsies, the detection and analysis of circulating tumor cells and disseminated tumor cells by less-invasive sampling of BM and PB has been shown to be of clinical relevance in many cancer types.9Wang H. Stoecklein N.H. Lin P.P. Gires O. Circulating and disseminated tumor cells: diagnostic tools and therapeutic targets in motion.Oncotarget. 2017; 8: 1884-1912Crossref PubMed Scopus (39) Google Scholar To improve the outcome of high-risk NB patients, the accurate and sensitive detection of MRD is essential to monitor the disease burden and treatment response. Because no recurrent oncogenic-fusion gene is identified in NB cells, MRD has been identified by detecting NB-associated mRNAs (NB-mRNAs) with real-time quantitative PCR (qPCR), which can define a cut-off value between NB cells and normal BM or PB cells.10Beiske K. Ambros P.F. Burchill S.A. Cheung I.Y. Swerts K. Detecting minimal residual disease in neuroblastoma patients—the present state of the art.Cancer Lett. 2005; 228: 229-240Crossref PubMed Scopus (38) Google Scholar,11Brownhill S.C. Burchill S.A. PCR-based amplification of circulating RNAs as prognostic and predictive biomarkers—focus on neuroblastoma.Pract Lab Med. 2017; 7: 41-44Crossref PubMed Scopus (5) Google Scholar Although single NB-mRNA such as tyrosine hydroxylase (TH) and paired-like homeobox 2b were in BM and PB S.A. B. Selby P. Neuroblastoma cell detection by tyrosine hydroxylase J Cancer. PubMed Scopus Google A. R. M. R. is a and minimal residual disease in Clin Oncol. PubMed Scopus Google Scholar NB-mRNAs were shown to be to tumor and sensitive MRD several sets of NB-mRNAs have been to have a prognostic value NB R. prognostic value of of marrow disease in patients over with J Cancer. Full Text Full Text PDF PubMed Scopus (38) Google Scholar, A. Swerts K. A. S. P. R. D. G. R. Burchill S.A. Neuroblastoma mRNAs outcome in children with a Clin Oncol. PubMed Scopus Google Scholar, D. Cheung I.Y. marrow minimal residual disease was an and a of survival after Clin Oncol. PubMed Scopus Google Scholar, A. B. H. R. N. S. S. R. K. M. Matthay K.K. S. S. R. of neuroblastoma in bone marrow or blood of patients with neuroblastoma provides a new disease and Res. 2017; PubMed Scopus Google Scholar However, NB-mRNAs by centers and to be in the NB-mRNAs homeobox and TH mRNAs) on in NB cancer stem cells by A. Hasegawa D. Yamamoto N. A. Ishida T. K. Kosaka Y. H. Yamamoto T. S. S. Mori T. T. A. Y. Iijima K. M. H. Nishimura N. Minimal residual disease in neuroblastoma patients on the of a set of real-time in PubMed Scopus Google Scholar This MRD in BM and PB has the detection of relapse/regrowth in two high-risk NB S. A. A. T. Hasegawa D. K. Kosaka Y. M. Yamamoto N. Mori T. A. Iijima K. H. Nishimura N. detection of tumor relapse/regrowth by minimal residual disease in high-risk neuroblastoma Lett. PubMed Scopus (10) Google Scholar In the present study, 7NB-mRNAs (CRMP1, DBH, DDC, GAP43, ISL1, PHOX2B, and TH mRNAs) digital PCR MRD in high-risk NB patients to more detection of of NB-mRNAs than high-risk NB patients, as defined by the Neuroblastoma S.L. T. Ambros P.F. G.M. A. B. T. D. T. A. Matthay K.K. Neuroblastoma an Clin Oncol. PubMed Scopus Google Scholar were at or between 2011 and on the or of the Japanese Neuroblastoma T. K. M. T. H. T. A. T. A. T. S. H. T. T. M. H. T. A. of a high-risk neuroblastoma treatment a from the Neuroblastoma J Clin Oncol. 2018; PubMed Scopus Google Scholar were BM and PB samples with were collected as as the of high-risk NB This study was by the at of and and was in accordance with the of of was at BM and PB sampling time point and BM on the and were combined into an H. Ambros B. Matthay K.K. neuroblastoma PubMed Scopus Google Scholar was in accordance with the Neuroblastoma G.M. B. M. of the neuroblastoma and to Clin Oncol. PubMed Scopus Google S.A. K. H. Ambros P.F. R. M. the of bone marrow disease and in children with neuroblastoma on of the Neuroblastoma 2017; PubMed Scopus Google Scholar and the was which to or stable, which to or no or which to patients with collection time the most disease status was a new disease status BM and PB samples were to stable, or to BM and response. BM samples from were from from samples from 56 samples from samples from and samples from PB samples from were from samples were from the Japanese of blood was by the and the Japanese BM and PB samples from NB patients and were using and cells were collected to the BM cells from cells were to the was with a to the and at with a and its using a to the was from or using a that and of and and to a of or samples were at after the BM and PB samples. ddPCR was using a ddPCR in a of 20 of ddPCR each of and of and to with a Droplet each was with a were a at by of and and a at was ddPCR were with a Droplet and the was using using were set as and To in the of and the of the was using as an that in a of of was used to the reproducible ddPCR and was two to to a of ddPCR were two to samples from NB patients and ddPCR analysis was in accordance with the digital the of PCR K. R. M. T. S.A. digital of quantitative digital PCR PubMed Scopus Google Scholar qPCR was using an PCR in a of of of each of and and of the to of was with a at by at and and at and was in To in the of and in the of and were used as was by the as A. B. H. R. N. S. S. R. K. M. Matthay K.K. S. S. R. of neuroblastoma in bone marrow or blood of patients with neuroblastoma provides a new disease and Res. 2017; PubMed Scopus Google Scholar the value each gene was set as the the amplification a of over and was this was not by the the detection were as the of the the detection the of the of the a higher level of qPCR analysis was in accordance with the S.A. M. R. T. of quantitative real-time PCR PubMed Scopus Google Scholar and were using at the were used ddPCR and qPCR from the and and and and and and TH and and and and and and were used TH and level of each NB-mRNA was defined as the of each NB-mRNA NB-mRNA by and by level of 7NB-mRNAs was defined as the of of each of the in samples was used each the of levels between BM and the level of 7NB-mRNAs in BM was as the of and samples. a between ddPCR and the level of 7NB-mRNAs was defined as the of of each in the levels of each NB-mRNA and 7NB-mRNAs between two were by the of the level of 7NB-mRNAs between BM and PB samples was by the diagnostic and prognostic of the levels of each NB-mRNA and 7NB-mRNAs was using G. a evaluation in clinical PubMed Scopus Google Scholar were two and was were with and which is a of to used in Y. of the 48: PubMed Scopus Google Scholar of 20 high-risk NB patients at or were in the present study, with a of at to and a time of to 73 20 patients, of tumor of had and of were BM at high-risk NB treatment that was used in the study BM and PB were as as BM samples were collected from the and S.A. K. H. Ambros P.F. R. M. the of bone marrow disease and in children with neuroblastoma on of the Neuroblastoma 2017; PubMed Scopus Google Scholar time were as treatment including high-dose chemotherapy with autologous peripheral blood stem cell rescue, surgery, and post-treatment including and this in samples, treatment samples, post-treatment samples, and samples disease status at sampling time was retrospectively as stable, or by the in samples, samples, and samples of BM and PB samples were from as and In the present study, a of BM and 67 samples from 20 NB patients and BM and samples from were by at was or and was to the Neuroblastoma marrow at marrow at was or and was to the Neuroblastoma was or and was to the Neuroblastoma H. Ambros B. Matthay K.K. neuroblastoma PubMed Scopus Google marrow at was or and was to the Neuroblastoma G.M. B. M. of the neuroblastoma and to Clin Oncol. PubMed Scopus Google S.A. K. H. Ambros P.F. R. M. the of bone marrow disease and in children with neuroblastoma on of the Neuroblastoma 2017; PubMed Scopus Google Scholar in a new time collection time high-dose chemotherapy with autologous peripheral blood stem cell surgery, and collection time from the of treatment to the clinical of bone peripheral collection time high-dose chemotherapy with autologous peripheral blood stem cell surgery, and collection time from the of treatment to the clinical of in a new bone peripheral MRD detection of relapse/regrowth by MRD in two high-risk NB A. Hasegawa D. Yamamoto N. A. Ishida T. K. Kosaka Y. H. Yamamoto T. S. S. Mori T. T. A. Y. Iijima K. M. H. Nishimura N. Minimal residual disease in neuroblastoma patients on the of a set of real-time in PubMed Scopus Google S. A. A. T. Hasegawa D. K. Kosaka Y. M. Yamamoto N. Mori T. A. Iijima K. H. Nishimura N. detection of tumor relapse/regrowth by minimal residual disease in high-risk neuroblastoma Lett. PubMed Scopus (10) Google Scholar To the more detection of low levels of NB-mRNA than MRD DBH, DDC, GAP43, ISL1, PHOX2B, and TH mRNAs were used MRD in the present Although NB-mRNAs were no or in normal cells by ddPCR in to and in to of BM and PB samples, level of each NB-mRNA in BM and PB varied between and collection time and was significantly higher in samples BM and PB samples collected from NB patients at than samples BM and PB samples from of NB-mRNAs in and as and as samples the of samples whose level of each NB-mRNA was than of and as and as samples the of samples whose level of each NB-mRNA was than of bone DBH, DDC, GAP43, ISL1, homeobox neuroblastoma-associated peripheral PHOX2B, paired-like homeobox tyrosine and as samples the of samples whose level of each NB-mRNA was than of samples. in a new bone DBH, DDC, GAP43, ISL1, homeobox neuroblastoma-associated peripheral PHOX2B, paired-like homeobox tyrosine MRD was as the of or two of NB-mRNAs a cut-off value between NB cells and normal BM or PB R. prognostic value of of marrow disease in patients over with J Cancer. Full Text Full Text PDF PubMed Scopus (38) Google Scholar, A. Swerts K. A. S. P. R. D. G. R. Burchill S.A. Neuroblastoma mRNAs outcome in children with a Clin Oncol. PubMed Scopus Google Scholar, D. Cheung I.Y. marrow minimal residual disease was an and a of survival after Clin Oncol. PubMed Scopus Google A. Hasegawa D. Yamamoto N. A. Ishida T. K. Kosaka Y. H. Yamamoto T. S. S. Mori T. T. A. Y. Iijima K. M. H. Nishimura N. Minimal residual disease in neuroblastoma patients on the of a set of real-time in PubMed Scopus Google Scholar However, the combined signature of NB-mRNAs was shown to have a higher diagnostic value than other NB-mRNA in MRD by A. B. H. R. N. S. S. R. K. M. Matthay K.K. S. S. R. of neuroblastoma in bone marrow or blood of patients with neuroblastoma provides a new disease and Res. 2017; PubMed Scopus Google Matthay K.K. R. S.A. A. Cohn S.L. J.M. peripheral blood high-risk neuroblastoma a Oncol. Full Text Full Text PDF PubMed Scopus Google Scholar To whether the combined signature of NB-mRNAs was to each NB-mRNA signature in MRD by analysis was the level of each NB-mRNA and defined as the of the level of each in samples BM and and samples BM and with MRD A. B. H. R. N. S. S. R. K. M. Matthay K.K. S. S. R. of neuroblastoma in bone marrow or blood of patients with neuroblastoma provides a new disease and Res. 2017; PubMed Scopus Google Scholar the level of 7NB-mRNAs was as a higher the than the level of each NB-mRNA the level of 7NB-mRNAs in a between the and samples and was used in the of NB-mRNAs in signature is as the TH signature is as the of DBH, DDC, GAP43, ISL1, PHOX2B, and TH neuroblastoma-associated bone DBH, DDC, GAP43, ISL1, homeobox peripheral PHOX2B, paired-like homeobox tyrosine signature is as the signature is as the of in a new DBH, DDC, GAP43, ISL1, PHOX2B, and TH neuroblastoma-associated bone DBH, DDC, GAP43, ISL1, homeobox peripheral PHOX2B, paired-like homeobox tyrosine Because the of between BM and PB was in MRD A. B. H. R. N. S. S. R. K. M. Matthay K.K. S. S. R. of neuroblastoma in bone marrow or blood of patients with neuroblastoma provides a new disease and Res. 2017; PubMed Scopus Google Scholar the of the level of 7NB-mRNAs between BM and PB was in the present MRD 208 BM and 67 PB samples, of BM and PB samples were collected at time treatment time post-treatment time and time level of 7NB-mRNAs in samples a between BM and PB but was higher in BM than PB To the level of 7NB-mRNAs in to disease burden, disease status was BM and PB samples, and was as stable, or the and level of 7NB-mRNAs in BM and PB samples with disease In it significantly in of disease status stable, or progression) the In it significantly in other than the and of 7NB-mRNAs in NB 208 67 and as and as and as and as status time point samples collected high-dose chemotherapy with autologous peripheral blood stem cell surgery, and samples collected from the of treatment to the clinical of and as and as and as and as time point samples collected high-dose chemotherapy with autologous peripheral blood stem cell surgery, and samples collected from the of treatment to the clinical of DBH, DDC, GAP43, ISL1, PHOX2B, and TH neuroblastoma-associated bone DBH, DDC, GAP43, ISL1, homeobox peripheral PHOX2B, paired-like homeobox tyrosine and as samples collected high-dose chemotherapy with autologous peripheral blood stem cell surgery, and samples collected from the of treatment to the clinical of in a new DBH, DDC, GAP43, ISL1, PHOX2B, and TH neuroblastoma-associated bone DBH, DDC, GAP43, ISL1, homeobox peripheral PHOX2B, paired-like homeobox tyrosine of the level of 7NB-mRNAs to the collection time point was level of 7NB-mRNAs in BM and PB samples varied with treatment and disease In a was in of collection time treatment, post-treatment, and the In was shown in the or other than the Because the level of 7NB-mRNAs in BM significantly between post-treatment and samples its with outcome was and post-treatment BM samples were into nonrelapsed/nonregrown and relapsed/regrown samples of the sampling time from chemotherapy to and in treatment BM samples In samples, the level of 7NB-mRNAs in samples not significantly from 17 samples In 73 post-treatment samples, 17 samples a significantly higher level of 7NB-mRNAs than 56 samples a between the level of 7NB-mRNAs in BM and tumor Although ddPCR potentially provides more and reproducible detection of low levels of NB-mRNAs than prognostic have been in the same samples from NB patients. determined the level of each NB-mRNA (CRMP1, DBH, DDC, GAP43, ISL1, PHOX2B, and TH in the same 73 and 17 post-treatment BM samples by qPCR was level of qPCR 7NB-mRNAs A. B. H. R. N. S. S. R. K. M. Matthay K.K. S. S. R. of neuroblastoma in bone marrow or blood of patients with neuroblastoma provides a new disease and Res. 2017; PubMed Scopus Google Scholar level of qPCR 7NB-mRNAs value was and the was To a between qPCR and the was the level of ddPCR 7NB-mRNAs in the same 73 and 17 post-treatment BM samples level of ddPCR 7NB-mRNAs a value of and an of with that ddPCR can a better prognostic value than qPCR in detecting MRD in NB samples. level of ddPCR 7NB-mRNAs in the of value of and of with a better prognostic value of ddPCR. These that a level of ddPCR 7NB-mRNAs in BM was associated with tumor relapse/regrowth in high-risk NB patients. In the present study, determined the level of DBH, DDC, GAP43, ISL1, PHOX2B, and TH in BM and PB by ddPCR MRD in high-risk NB patients. To this is the study of MRD in NB patients and that ddPCR detection of 7NB-mRNAs in BM has a prognostic value in tumor relapse/regrowth of high-risk NB patients ddPCR is the most sensitive to the of and potentially provides more and in NB-mRNAs with P.J. E. of targets PCR by of Google M. by digital PCR real-time PubMed Scopus Google Scholar To the most of the of ddPCR over D. M. and of by digital Full Text Full Text PDF PubMed Scopus Google M. S. K. C. T. S. M. H. and a of the evaluation of residual disease in 2017; PubMed Scopus Google Scholar was as an NB-mRNA and the was to to of each NB-mRNA was in more than and of BM and PB samples, ddPCR detection of low levels of NB-mRNA in more than samples the time that was no in the of NB-mRNAs between and samples and This to the cut-off value MRD evaluation in NB samples. and set a cut-off value each NB-mRNA on its in normal cells, and samples as MRD the of or two NB-mRNA the cut-off A. Hasegawa D. Yamamoto N. A. Ishida T. K. Kosaka Y. H. Yamamoto T. S. S. Mori T. T. A. Y. Iijima K. M. H. Nishimura N. Minimal residual disease in neuroblastoma patients on the of a set of real-time in PubMed Scopus Google I.Y. Y. Cheung gene to minimal residual disease of Res. PubMed Scopus Google A. B. R. R. Detecting minimal residual disease in the of a of real-time quantitative PCR PubMed Scopus Google Scholar However, a study that the combined signature of NB-mRNAs had a better diagnostic value than other NB-mRNA in MRD by A. B. H. R. N. S. S. R. K. M. Matthay K.K. S. S. R. of neuroblastoma in bone marrow or blood of patients with neuroblastoma provides a new disease and Res. 2017; PubMed Scopus Google Scholar In the present study, that the level of 7NB-mRNAs higher than the level of each NB-mRNA by ddPCR ddPCR potentially provides more and reproducible detection of low levels of NB-mRNAs than has not been present study whether ddPCR detection of 7NB-mRNAs tumor relapse/regrowth more than qPCR detection of the same 7NB-mRNAs in the same post-treatment BM samples high-risk NB patients. present have shown the time that ddPCR detection of 7NB-mRNAs a higher than qPCR detection of 7NB-mRNAs It a of ddPCR detection of 7NB-mRNAs in a clinical Although the of NB-mRNA between BM and PB by qPCR has A. B. H. R. N. S. S. R. K. M. Matthay K.K. S. S. R. of neuroblastoma in bone marrow or blood of patients with neuroblastoma provides a new disease and Res. 2017; PubMed Scopus Google A. B. R. R. Detecting minimal residual disease in the of a of real-time quantitative PCR PubMed Scopus Google Scholar the present MRD its with qPCR detection of DDC, PHOX2B, and TH in high-risk NB patients with A. B. H. R. N. S. S. R. K. M. Matthay K.K. S. S. R. of neuroblastoma in bone marrow or blood of patients with neuroblastoma provides a new disease and Res. 2017; PubMed Scopus Google Scholar ddPCR detection of 7NB-mRNAs (CRMP1, DBH, DDC, GAP43, ISL1, PHOX2B, and TH a between BM and but was higher in BM than PB in high-risk NB patients Although PB sampling was invasive and to the of than BM E. P. Circulating tumor as a PubMed Scopus Google Scholar PB was not a sensitive BM in NB patients. most of the present study was the of ddPCR detection of 7NB-mRNAs with relapse/regrowth in post-treatment BM samples from high-risk NB patients and This that MRD in BM risk stratification of high-risk NB patients. other have shown that NB-mRNAs in BM and PB by qPCR a prognostic value high-risk NB R. prognostic value of of marrow disease in patients over with J Cancer. Full Text Full Text PDF PubMed Scopus (38) Google Scholar, A. Swerts K. A. S. P. R. D. G. R. Burchill S.A. Neuroblastoma mRNAs outcome in children with a Clin Oncol. PubMed Scopus Google Scholar, D. Cheung I.Y. marrow minimal residual disease was an and a of survival after Clin Oncol. PubMed Scopus Google Scholar, A. B. H. R. N. S. S. R. K. M. Matthay K.K. S. S. R. of neuroblastoma in bone marrow or blood of patients with neuroblastoma provides a new disease and Res. 2017; PubMed Scopus Google Scholar of levels of DDC, GAP43, PHOX2B, and TH and at the of in BM was associated with a R. prognostic value of of marrow disease in patients over with J Cancer. Full Text Full Text PDF PubMed Scopus (38) Google Scholar the of levels of PHOX2B, and TH was associated with a outcome at in BM or PB and at the of in A. Swerts K. A. S. P. R. D. G. R. Burchill S.A. Neuroblastoma mRNAs outcome in children with a Clin Oncol. PubMed Scopus Google Scholar the more of NB patients, survival of NB patients was associated with the level of DDC, PHOX2B, and TH A. B. H. R. N. S. S. R. K. M. Matthay K.K. S. S. R. of neuroblastoma in bone marrow or blood of patients with neuroblastoma provides a new disease and Res. 2017; PubMed Scopus Google Scholar and survival of NB patients with was by the level of ISL1, and after two of immunotherapy in D. Cheung I.Y. marrow minimal residual disease was an and a of survival after Clin Oncol. PubMed Scopus Google Scholar However, were several in the present Although detection of 7NB-mRNAs (CRMP1, DBH, DDC, GAP43, ISL1, PHOX2B, and TH in BM was associated with tumor relapse/regrowth and correlated significantly with that in its prognostic value in PB has the level of 7NB-mRNAs at in BM not tumor relapse/regrowth in to the predictive of PHOX2B, and TH at A. Swerts K. A. S. P. R. D. G. R. Burchill S.A. Neuroblastoma mRNAs outcome in children with a Clin Oncol. PubMed Scopus Google Scholar These be from the of NB patients and in the present In ddPCR is a and reproducible to low levels of NB-mRNAs in BM and PB samples collected from high-risk NB patients. levels of 7NB-mRNAs in BM by ddPCR tumor relapse/regrowth in high-risk NB patients. MRD improve detection of evaluation of disease status and treatment and of in high-risk NB patients. clinical study ddPCR detection of 7NB-mRNAs in the of in of NB patients and be to its clinical the at and the and at BM and PB samples. with with with