[Establishment of a preclinical neuroblastoma model in immunocompetent mice]

To develop a NB animal model which makes possible studies related to tumor immunity.

Two types of NB cells were used. Cell line 36769 was derived from TH-MYCN+ mouse in which overexpression of the MYCN gene is governed by rat tyrosine hydroxylase promotor. Cell line 4040 was derived from TH-MYCN/ALK mice, which in addition express an activating mutation of ALK gene. For each cell type, 1x10⁶ neurospheres were implanted in 129/SVJ mice (with the same genetic background as donors, n=8), via orthotopic injection in the left suprarenal gland by intraperitoneal approach, through a transverse supraumbilical laparotomy. Daily postsurgical clinical follow-up of the animals was done until they were sacrificed at four weeks. The tumor presence was macroscopically confirmed. The tumoral sample was excised and was processed for cellular immunity and molecular tolerance mediator's studies. The existence of metastasis was investigated by flow cytometry in the spleen, bone marrow and peripheral blood.

1) Orthotopic Neuroblastoma was generated in all the transplanted mice. 2) The tumors were infiltrated by several immune subpopulations, with effector, regulatory and suppressor inmunophenotype. This was similar to the inmunophenotype described in human NB. Furthermore, the molecular mediators of the environment point to a state of protumoral tolerance.

The orthotopic implantation of NB neurospheres in syngeneic mice has allowed us to generate a NB model in which it has been possible to study the tumor immunity.