RESUMEN
There are considerable differences in tumour biology between adult and paediatric cancers. The existence of cancer initiating cells/cancer stem cells (CIC/CSC) in paediatric solid tumours is currently unclear. Here, we show the successful propagation of primary human Wilms' tumour (WT), a common paediatric renal malignancy, in immunodeficient mice, demonstrating the presence of a population of highly proliferative CIC/CSCs capable of serial xenograft initiation. Cell sorting and limiting dilution transplantation analysis of xenograft cells identified WT CSCs that harbour a primitive undifferentiated-NCAM1 expressing-"blastema" phenotype, including a capacity to expand and differentiate into the mature renal-like cell types observed in the primary tumour. WT CSCs, which can be further enriched by aldehyde dehydrogenase activity, overexpressed renal stemness and genes linked to poor patient prognosis, showed preferential protein expression of phosphorylated PKB/Akt and strong reduction of the miR-200 family. Complete eradication of WT in multiple xenograft models was achieved with a human NCAM antibody drug conjugate. The existence of CIC/CSCs in WT provides new therapeutic targets.
Asunto(s)
Neoplasias Renales/metabolismo , Neoplasias Renales/patología , Células Madre Neoplásicas/patología , Tumor de Wilms/metabolismo , Tumor de Wilms/patología , Antígeno AC133 , Aldehído Deshidrogenasa/metabolismo , Familia de Aldehído Deshidrogenasa 1 , Animales , Anticuerpos Monoclonales/uso terapéutico , Antígenos CD/metabolismo , Antígeno CD56/metabolismo , Diferenciación Celular , Proliferación Celular , Separación Celular/métodos , Expresión Génica , Glicoproteínas/metabolismo , Humanos , Neoplasias Renales/genética , Neoplasias Renales/terapia , Maitansina/análogos & derivados , Maitansina/uso terapéutico , Ratones , Ratones Endogámicos NOD , Ratones SCID , Células Madre Neoplásicas/metabolismo , Péptidos/metabolismo , Retinal-Deshidrogenasa , Células Tumorales Cultivadas , Ensayo de Tumor de Célula Madre , Tumor de Wilms/genética , Tumor de Wilms/terapia , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
During development, renal stem cells reside in the nephrogenic blastema. Wilms' tumour (WT), a common childhood malignancy, is suggested to arise from the nephrogenic blastema that undergoes partial differentiation and as such is an attractive model to study renal stem cells leading to cancer initiation and maintenance. Previously we have made use of blastema-enriched WT stem-like xenografts propagated in vivo to define a 'WT-stem' signature set, which includes cell surface markers convenient for cell isolation (frizzled homolog 2 [Drosophila] - FZD2, FZD7, G-protein coupled receptor 39, activin receptor type 2B, neural cell adhesion molecule - NCAM). We show by fluorescenceactivated cell sorting analysis of sphere-forming heterogeneous primary WT cultures that most of these markers and other stem cell surface antigens (haematopoietic, CD133, CD34, c-Kit; mesenchymal, CD105, CD90, CD44; cancer, CD133, MDR1; hESC, CD24 and putative renal, cadherin 11), are expressed in WT cell sub-populations in varying levels. Of all markers, NCAM, CD133 and FZD7 were constantly detected in low-to-moderate portions likely to contain the stem cell fraction. Sorting according to FZD7 resulted in extensive cell death, while sorted NCAM and CD133 cell fractions were subjected to clonogenicity assays and quantitative RT-PCR analysis, exclusively demonstrating the NCAM fraction as highly clonogenic, overexpressing the WT 'stemness' genes and topoisomerase2A (TOP2A), a bad prognostic marker for WT. Moreover, treatment of WT cells with the topoisomerase inhibitors, Etoposide and Irinotecan resulted in down-regulation of TOP2A along with NCAM and WT1. Thus, we suggest NCAM as a marker for the WT progenitor cell population. These findings provide novel insights into the cellular hierarchy of WT, having possible implications for future therapeutic options.