Discrepant flow cytometric expression and function of P-glycoprotein in neuroblastic tumors

Patients suffering from neuroblastic tumors are currently being classified into prognostic subsets based on different clinical and biologic features. In this study, a triple-color flow cytometric assay and a functional test were applied to neuroblastoma cell lines and patients with a neuroblastic tumor, and the value of P-glycoprotein expression and function as potential prognostic characteristics, was determined.

Twenty-two single-cell suspensions prepared from tumors, and neuroblasts from four bone marrow samples were analyzed by triple-color flow cytometry. Neuroblasts were identified by NB84-positivity and absence of CD45. P-glycoprotein expression was evaluated using 4E3 and MRK16 antibodies. Eighteen samples were tested with a functional assay, based on accumulation and retention of rhodamine-123 with and without the inhibitor verapamil. Six neuroblastoma cell lines were also evaluated.

P-glycoprotein expression was seen in 18 of 26 patient samples and in three of six cell lines. The highest expression levels were found in low stage neuroblastoma and well-differentiated tumors; whereas the highest activities were found in stage 4 neuroblastoma and the lowest in ganglioneuroblastoma and ganglioneuroma patients. In 10 of 17 samples, concordant results were found between the flow cytometric immunological test and immunocytochemistry.

The described flow cytometric technique is a new, alternative approach to detect P-glycoprotein expression and function in neural crest tumors. Based on the expression level and the activity value, patients can be segregated into different phenotypic groups. In particular, those patients with high P-glycoprotein activity might benefit from treatment regimens containing reversal agents.