Abstract 1626: Evaluation of the expression of neuroblastoma-associated genes for bone marrow (BM) involvement and minimal residual disease (MRD) detection
Authors: Druy Alexander E. , Shorikov Egor V. , Tsaur Grigory A. , Popov Alexander M. , Saveliev Leonid I. , Fechina Larisa G. 
Published: August 2015

Methods. Panel of 4 tumor-associated genes (TAGs): PHOX2B, TH, ELAVL4 and GD2 was created. Normalized expression (CtABL-CtTAG) of these genes was analyzed in neuroblastoma (NB) cell lines, in 26 BM samples from patients without malignancies, in the discovery (331 BM samples from 57 patients) and validation cohorts of NB patients (311 BM samples from 55 patients) and in 23 peripheral blood stem cells (PBSC) preparations. For performing of ROC analysis BM samples were defined as positive in case of detectable PHOX2B expression or tumor cells presence in BM smears. Threshold levels (TL) of each gene expression were established and subsequently applied for overall correct prediction (OCP) computation. Prognostic significance of the BM positivity for the evaluated markers was determined using event-free (EFS) and overall survival (OS) rates with median of follow-up time 2.45 years.

Results. Neither PHOX2B nor TH expression was detected in the intact BM, while expression of ELAVL4 was revealed in 20 and GD2 - in 15 of 26 BM samples of patients without malignancies. In the discovery cohort 105 analyzed BM samples were positive for PHOX2B expression, while 101 for TH. TH expression was revealed in 5 out of 224 negative BM samples. ELAVL4 and GD2 expression was detected in all 107 positive BM samples as well as in the majority of negative BM samples (209 and 197 of 224, respectively). TL of TH, ELAVL4 and GD2 expression established by ROC analysis were -16.535, -7.130 and -8.459. OCP values calculated using TL achieved 0.952, 0.828 and 0.767 correspondingly. OCP for PHOX2B was 0.994. Due to high OCP values for PHOX2B and TH expression of these genes was analyzed in the validation cohort, where TL for TH was -13.995, while OCP values achieved 0.997 for PHOX2B and 0.939 for TH.

Presence of PHOX2B/TH expression in BM at the time of primary NB diagnostics led to decreased both EFS (.31SE.12 vs .81SE.06, p&lt;.001) and OS (.31SE.13 vs .87SE.05, p&lt;.001). Persistence of TAGs expression during the treatment demonstrated trend to reduced EFS (.27SE.12 vs .43SE.19, p = .08). No expression of TAGs was find in the PBSC preparations while positivity for PHOX2B/TH expression before PBSC apheresis had strongly negative prognostic impact (EFS .00 vs .35SE.14, p = .04; OS .00 vs .36SE.15, p = .03) despite of CD34+ selection. Predominance of PHOX2B expression over TH in BM of primary NB patients with the difference in normalized expression greater then 1.68 had negative prognostic significance: EFS .00 vs .56SE.12, p = .017, OS .00 vs .72SE.11, p = .006.

Conclusion. PHOX2B and TH are the most appropriate markers for BM involvement and MRD detection in NB patients. Presence of these TAGs expression in the BM at the time of primary diagnostics and before PBSC apheresis had strongly negative prognostic impact. Predominance of PHOX2B expression over TH in the BM can define high risk patients.

Citation Format: Alexander E. Druy, Egor V. Shorikov, Grigory A. Tsaur, Alexander M. Popov, Leonid I. Saveliev, Larisa G. Fechina. Evaluation of the expression of neuroblastoma-associated genes for bone marrow (BM) involvement and minimal residual disease (MRD) detection. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1626. doi:10.1158/1538-7445.AM2015-1626